# 705: BACILLUS INFERNUS MEDIUM


## Main sol. 705

5.3	g	Ferric citrate monohydrate	
5.8	g	NaCl	
1	g	NH4Cl	
0.4	g	K2HPO4 x 3 H2O	
1	g	MgCl2 x 6 H2O	
2	g	Yeast extract (OXOID)	
2	g	Trypticase peptone (BD BBL)	
10	ml	**Trace element solution**	
0.8	g	Na-acetate	
1.2	g	Na-lactate	
0.5	g	2-Mercaptoethanesulfonate (coenzyme M)	
1000	ml	Distilled water	

1. First, dissolve ferric citrate in water by heating and adjust to pH 7.5 with NaOH, then add other
medium ingredients except acetate, lactate, and coenzyme M. Sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add acetate, lactate, and coenzyme M from sterile anoxic stock solutions prepared under 100% N2 gas.
2. Prior to inoculation the pH of the complete medium should be checked and adjusted to 7.5, if necessary.


## Trace element solution

0.5	g	Na2-EDTA x 2 H2O	
0.15	g	CoCl2 x 6 H2O	
0.1	g	MnCl2 x 4 H2O	
0.1	g	FeSO4 x 7 H2O	
0.1	g	ZnCl2	
40	mg	AlCl3 x 6 H2O	
40	mg	Na2WO4 x 2 H2O	
30	mg	Na2SeO3 x 5 H2O	
20	mg	NiCl2 x 6 H2O	
20	mg	CuCl2 x 2 H2O	
10	mg	H3BO3	
10	mg	Na2MoO4 x 2 H2O	
1000	ml	Distilled water	

1. First dissolve EDTA in distilled water and adjust pH to 7 using 2 N NaOH, then dissolve remaining compounds.