For the beta-galactosidase kinetic assay, 100 mL of a 275 mM sodium phosphate bu
ID: 1023430 • Letter: F
Question
For the beta-galactosidase kinetic assay, 100 mL of a 275 mM sodium phosphate buffer containing 2.75 mM MgCl_2 at pH 7.5 will be prepared. The following will be available: phosphoric acid (15 M), monosodium phosphate (FW 137.99), disodium phosphate (FW 174.20), trisodium phosphate (FW 380.12), and MgCl_2 (FW 203.30). Calculate the mass of reagents required to prepare 100 ml of a 275 mM solution of monobasic phosphate and 100 ml of a 275 mM solution of dibasic phosphate, each containing 2.75 mM MgCl_2. Calculate the volume of each of these solutions that will be combined to make 100 mL of a 275 mM sodium phosphate buffer containing 2.75 mM MgCl_2 at pH 7.5.Explanation / Answer
100 mL is 0.1 L
so 0.1 L of 275 mM solution means
0.1 x 275 x 10-3 M = 0.0275 moles
0.1 x 2.75 x 10-3 M = 0.000275 moles
0.000275 x 203.3 = 0.0559 g of MgCl2
Amount of monosodium phosphate = 0.0275 x 137.99 = 3.79 g dissolved in 100 mL with 0.0559 g of MgCl2
Amount of disodium phosphate = 0.0275 x 174.2 = 4.79 g dissolved in 100 mL with 0.0559 g of MgCl2
pH = pKa + log base/acid
7.5 = 7.21 + log base/acid
log base/acid = 0.29
base/acid = 100.29
base/acid = 1.95
SO we will take 66 mL of the disodium phosphate solution and 34 mL of the monosodium phosphate solution from above then the ratio will be 1.95 and it will have 2.75 mM MgCl2