Citric Acid – Na₂HPO₄ Buffer Preparation, pH 2.6–7.61

From Sigma Buffer Reference Center

Citric acid monohydrate, C₆H₈O₇ • H₂O,
MW 210.14;
0.1 M contains 21.01 g/L.

Na₂HPO₄, MW 141.98;
0.2 M contains 28.40 g/L, or

Na₂HPO₄ • 2H₂O, MW 178.05;
0.2 M contains 35.61 g/L.

pH	mL 0.1 M-citric acid	mL 0.2-Na2HPO4
2.6	89.10	10.90
2.8	84.15	15.85
3.0	79.45	20.55
3.2	75.30	24.70
3.4	71.50	28.50
3.6	67.80	32.20
3.8	64.50	35.50
4.0	61.45	38.55
4.2	58.60	41.40
4.4	55.90	44.10
4.6	53.25	46.75
4.8	50.70	49.30
5.0	48.50	51.50
5.2	46.40	53.60
5.4	44.25	55.75
5.6	42.00	58.00
5.8	39.55	60.45
6.0	36.85	63.15
6.2	33.90	66.10
6.4	30.75	69.25
6.6	27.25	72.75
6.8	22.75	77.25
7.0	17.65	82.35
7.2	13.05	86.95
7.4	9.15	90.85
7.6	6.35	93.65

For 2021 QBoC

Need pH 3,4,5,6,7

Start with

• Citric acid (100 mM) (borrowed from P-Chem) 2.65 g
  • 125 mL H2O
• Sodium phosphate dibasic (200 mM)
  • MW = 268.07
  • 200 mM = 8.042 g
  • 150 mL H2O

Then make

pH	x mL 0.1 M citric acid	y mL 0.2M Na2HPO4
3.0	39.725	10.275
4.0	30.75	19.275
5.0	24.25	25.75
6.0	18.425	31.575
7.0	8.825	41.175

Test pH of 1:10, 1:2

• pH remains constant when diluted to 20% with LB.
• So make 50 mL of each diluted to 40% with LB. Students will dilute it 1:1 with LB
  • 20 mL buffer
  • 30 mL water
• Filter steriize, aliquot for students