Question

You are given the following: water, 10 mM phosphate buffer pH 7.4, EDTA solid (FW 292.25 g/mol), 0.5 M Tris HCl, 3 M NaCl, OAA solid (FW 131.10), NADH solid (FW 709.40), and solutions of NaOH and HCl. Calculate and describe how you would prepare of each of the following:  

a. 50 mL of 0.5 M EDTA pH 8.0

b. 10 ml solution containing 10 mM Tris-Cl pH 8.0 + 100 mM NaCl

c. 10 mL of 20 mM OAA dissolved in 10 mM phosphate buffer pH 7.4

d. 10 mL of 2 mM NADH dissolved in 10mM phosphate buffer pH 7.4

Answer 1

a. Required volume* required concentration* molecular weight of EDTA = 0.05*0.5*292.25 = 7.31

So, we will add 7.31gm of EDTA in 30ml of distilled water. Then adjust the pH with NaOH to pH 8.0

b. Initial concentration*initial volume = final concentration*final volume

For Tris-Cl,

500mM* Xml = 10mM*10ml

X = 0.2ml

For NaCl,

3000mM* Xml = 100mM*10ml

X = 0.3ml

Total volume = 0.2+0.3 = 0.5ml

Finally,

Diluent = final volume - combined initial volume = 10ml - 0.5ml = 9.5 ml

So, we will add 9.5ml of distilled water.

c. For phosphate buffer preparation,

Solution A : 0.1M potassium dihydrogen phosphate

Dissolve 13.61gm KH₂PO₄ in 1000ml distilled water

Solution B : 0.1M disodium hydrogen phosphate

Dissolve 17.8gm Na₂HPO₄.2H₂O in 1000ml distilled water

For pH 7.4, take 20ml of solution A and 80ml of solution B.

For 10ml of 20mM OAA

0.01*0.02*131.10 = 2.62

So, we will add 2.62gm of OAA in 10ml of 10mM phosphate buffer.

D. For NaDH,

0.01*0.02*709.40 = 0.14

So, we will add 0.14gm of NaDH in 10ml of 10mM phosphate buffer.