8/12/2019 Acetate and Phosphate Buffers

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PHOSPHATE BUFFERS

Phosphate buffer (sodium salt) is typically prepared by mixing solutions of sodium phosphate dibasic

(Na2HPO4) and sodium phosphate monobasic (NaH2PO4).

The phosphate group is a coalent unit (i.e.! the Ps and Os are coalently bound to each other). The Na and H

atoms dissociate! and the H contributes the pH of the solution.

The monobasic form has more H" to contribute (per PO4unit) than the dibasic! and a solution of monobasic

#ill be slightly acidic (a $% solution has a pH of about 4.2) compared to the dibasic (#ith only one H" perPO4! a $% solution has a pH of about &.').

The monobasic! more acidicSolutionA is mixed #ith the dibasic! more basicSolutionB to get the desired

pH.

f the concentration of PO4is the same in both solutions! then the resulting solution #ill hae the same

concentration (of PO4) regardless of the proportions of each #hich are added! because $ ml of *.2 m PO4 " &$ml of *.2 + PO4, '** ml of *.2 + PO4-

Sodium phosphate buffer, 0.1 M

Solution A: 2./ g NaH2PO4H2O per liter (*.2 +)

Solution B: $0./$ g Na2HPO4H2O per liter (*.2 +)

1eferring to Table .2.0 for desired pH! mix the indicated olumes of 3olutions and ! thendilute #ith H2O to 2** ml

Potassium phosphate buffer, 0.1 M

Solution A: 2.2 g 5H2PO4per liter (*.2 +)

Solution B: 04.6 g 52HPO4per liter (*.2 +)

1eferring to Table .2.0 for desired pH! mix the indicated olumes of 3olutions and ! thendilute #ith H2O to 2** ml

This may be made as a 5 or 10-fold concentrate by scaling up the amount of potassium or sodiumphosphate in the same volume. hosphate buffers sho! concentration-dependent p" changes# so chec$concentrate p" by diluting an ali%uot to the final concentration .

Table A.2.3 Preparation of *.' + 3odium and Potassium Phosphate uffers

7esiredpH

3olution (ml)

3olution (ml)

7esiredpH

3olution (ml)

3olution (ml)

$. &0.$ /.$ /.& 4$.* $$.*$.6 &2.* 6.* .* 0&.* /'.*$.& &*.* '*.* .' 00.* /.*/.* 6. '2.0 .2 26.* 2.*

/.' 6$.* '$.* .0 20.* .*/.2 6'.$ '6.$ .4 '&.* 6'.*/.0 .$ 22.$ .$ '/.* 64.*/.4 0.$ 2/.$ ./ '0.* 6.*/.$ /6.$ 0'.$ . '*.$ &*.$/./ /2.$ 0.$ .6 6.$ &'.$/. $/.$ 40.$ .& .* &0.*/.6 $'.* 4&.* 6.* $.0 &4.

ACETATE BUFFERS

8/12/2019 Acetate and Phosphate Buffers

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cetate buffers are prepared by mixing e8uimolar solutions of acidic and basic forms of acetate in order to achieethe desired pH. The acidic form is acetic acid (HOc! or Hc)! and the basic form is the sodium or potassium salt(really the unprotonated Oc9). :*.2 + acetate; buffer contains a total of *.2 mole of :acetate; per liter!#hether it is protonated or not.

Potassium acetate buffer, 0.1 M

Solution A: ''.$$ ml glacial acetic acidou #ill see protocols for solutions of high concentration #hich seem li?e they are prepared differently 99 but ifyou loo? closely you #ill see that the same principles apply the concentration of the buffering ion (here! acetate) is#hat gies the solution its name! and as long as the pH is ad@usted #ith either the same concentration of that

buffering ion (a)! or the appropriate acid or base lac?ing that ion altogether (b)! the correct concentration #ill bemaintained.

(a) 3M Na Acetate, pH 5. (b) 5 M ! Acetate, pH ".#4*6 g Na acetate 0H2O in 6** ml of H2O 2&.$ ml glacial acetic acid

dd H2O to ' liter 5OH pellets to 4.6

d@ust pH to $.2 #ith 0 + acetic acid H2O to '** ml