# 1095: NATRANAEROBIUS MEDIUM


## Main sol. 1095

100	g	NaCl	
0.2	g	KH2PO4	
0.1	g	MgCl2	
0.5	g	NH4Cl	
10	g	Yeast extract (OXOID)	
10	g	Tryptone (BD Bacto)	
1	ml	**Trace element solution SL-10**	
68	g	Na2CO3	
38	g	NaHCO3	
0.7	g	L-Cysteine HCl x H2O	
1	ml	**Wolin's vitamin solution (10x)**	
5	g	Sucrose	
1000	ml	Distilled water	

1. Dissolve ingredients (except carbonates, cysteine, sucrose and vitamins), then sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Add carbonate, hydrogencarbonate and cysteine and adjust pH to 8.5. Dispense medium under 100% N2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving add sucrose and vitamins from sterile anoxic stock solutions prepared under 100% N2 gas and sterilized by filtration.


## Trace element solution SL-10

10	ml	HCl (25%)	
1.5	g	FeCl2 x 4 H2O	
70	mg	ZnCl2	
100	mg	MnCl2 x 4 H2O	
6	mg	H3BO3	
190	mg	CoCl2 x 6 H2O	
2	mg	CuCl2 x 2 H2O	
24	mg	NiCl2 x 6 H2O	
36	mg	Na2MoO4 x 2 H2O	
990	ml	Distilled water	

1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.


## Wolin's vitamin solution (10x)

20	mg	Biotin	
20	mg	Folic acid	
100	mg	Pyridoxine hydrochloride	
50	mg	Thiamine HCl	
50	mg	Riboflavin	
50	mg	Nicotinic acid	
50	mg	Calcium D-(+)-pantothenate	
1	mg	Vitamin B12	
50	mg	p-Aminobenzoic acid	
50	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water