Basic Concepts in Clinical Biochemistry-A Practical Guide.7z

aSince 1000 ml of 0.2 M acetic acid contain¼0.2 moles,

so 46 ml of 0.2 M acetic acid will contain¼0.246/1000¼ 9200 μmoles
Calculate pH using Henderson-Hasselbalch equation

pH¼pKaþlog

½Šconjugate base

½Šconjugate acid

pH¼ 4 : 86 þlog800= 9200

¼ 4 : 86  1 : 06 ¼ 3 : 8

6.6.2 0.2 M Sodium Phosphate Buffer (pKa 6.86)

Sodium phosphate buffer is prepared by mixing sodium dihydrogen phosphate
(NaH 2 PO 4 ) and disodium hydrogen phosphate (Na 2 HPO 4 ). Prepare 0.2 M solutions
of monobasic and dibasic salts separately and mix dibasic solution in monobasic
solution as given in table.

S. no

Volume of

0.2 M

NaH 2 PO 4

taken

Amount of 0.2 M

acetic acid in used

volume (μmole)

Volume of

0.2 M

Na 2 HPO 4

taken

Amount of 0.2 M

Na 2 HPO 4 in used

volume (μmole)

Theoretical

pH

1. 46 9200 a 4 800 5.7a


2. 42 8400 8 1600 6.1


3. 38 7600 12 2400 6.3


4. 34 6800 16 3200 6.5


5. 30 6000 20 4000 6.6

aSince 1000 ml of 0.2 M NaH 2 PO 4 contain¼0.2 moles,

so 46 ml of 0.2 M NaH 2 PO 4 will contain¼0.2 x 46/1000¼ 9200 μmoles
Calculate pH using Henderson-Hasselbalch equation

pH¼pKaþlog

½Šconjugate base

½Šconjugate acid

pH¼ 4 : 86 þlog800= 9200

¼ 6 : 86  1 : 06 ¼ 5 : 7

6.6.3 0.2 M Tris-HCl Buffer (pKa 8.1)

Tris buffer is prepared by mixing appropriate volumes of Tris and HCl as given in
table.

26 6 Preparation of General Laboratory Solutions and Buffers