Q1)To melanic acid[100ml of .05M] ,1gm of Tris[hydroxymethyl]amino methane(m.wt=121,pKa=8.3) is added.?

Question

pka1 of melanic acid=2.7, pka2=5.7
1)how many mM of melanic acid & tris are present?
2)which pka will you see to calculate the pH?
3)what is the overall pH of the soln?

Q2)wat are the advantages of using absorption at 597/466 rather than just at 595 for measuring proteins in coomassie blue/brilliant blue reagent?

Q3)The pkas of slutamic acid are approx. 2,4,10. wat is the pH of .1! soln of monododium glutamate?

Q4).1ml of stock soln is diluted with 2.4ml of distilled water. 50microlitres of this soln id diluted with .4ml of distilled water. the overall dilution is? ( can be givin in a ratio)

please can someone help me on this ASAP
thanks a tonn !

please could you elaborate the answer.. i have very little idea about it

its perfect otherwise :)
thanks

Answer 1

1)how many mM of melanic acid & tris are present?

Molarity (M) is defined as (moles/1L) and then mM would be (mM/mL). You will assume that the 1 g Tris doesn't change volume, so calculate mM of melanic acid using 100ml of 0.05M and calculate mM of tris by using molecular wt. (grams per mole) and the 1 gram.

pH = pKa + log [A-]/[HA]. or. pH = pKa + log [salt]/[acid] (Henderson Hasselbach equation)

You would need to use 2nd pKa since the melanic soln is dissociated (water has pH 7).

Q2) The binding of the dye to protein shifts the peak absorbance of the dye. Unbound Coomassie Blue absorbs light maximally at a wavelength of 465 nm, while the absorption maximum is at 595 nm when the dye is bound to protein. The absorbance of light by the dye-protein complex at 595 nm is proportional to the amount of protein bound (over a limited range); i.e. there is a linear relationship between absorbance and the total protein concentration of the sample over a narrow range. From http://www.acad.carleton.edu/curricular/BIOL/resources/rlink/lab1p4.html

This means that you can subtract out unbound dye.

Q3) I assume you mean 0.1M monosodium glutamate. It is monosodium so use pKa 2 in Henderson Hasselbach

Q4) 2.4mL + 0.1 mL= 2.5mL sln (0.1mL stock/2.5mL sln x100) = 4%

Now use C1V1=C2V2 to solve for final dilution

Answer 2

Melanic Acid