# 875: DESULFACINUM HYDROTHERMALE MEDIUM


## Main sol. 875

950	ml	**Solution A**	
1	ml	**Solution B**	
30	ml	**Solution C**	
1	ml	**Solution D**	
10	ml	**Solution E**	
10	ml	**Solution F**	

1. Solution A is sparged with 80% N2 and 20% CO2 gas mixture to reach a pH below 6 (at least 30 - 45 min), then distributed under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclaved. Solutions B, E and F are autoclaved separately under 100% N2 gas atmosphere. Solution C is autoclaved under 80% N2 and 20% CO2 gas atmosphere. Solution D is prepared under 100% N2 gas atmosphere and sterilized by filtration. To complete the medium appropriate amounts of solutions B to F are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be 7.0 - 7.3.


## Solution A

26	g	NaCl	
5.6	g	MgCl2 x 6 H2O	
1.4	g	CaCl2 x 2 H2O	
6.8	g	MgSO4 x 7 H2O	
0.25	g	NH4Cl	
0.2	g	KH2PO4	
0.72	g	KCl	
0.5	ml	Sodium resazurin (0.1% w/v)	
950	ml	Distilled water	


## Solution B

1	ml	**Trace element solution SL-10**	


## Solution C

1.5	g	Na2CO3	
30	ml	Distilled water	


## Solution D

1	ml	**Seven vitamins solution**	


## Solution E

2.5	g	Na-DL-lactate	
10	ml	Distilled water	


## Solution F

0.4	g	Na2S x 9 H2O	
10	ml	Distilled water	


## 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.


## Seven vitamins solution

100	mg	Vitamin B12	
80	mg	p-Aminobenzoic acid	
20	mg	D-(+)-biotin	
200	mg	Nicotinic acid	
100	mg	Calcium pantothenate	
300	mg	Pyridoxine hydrochloride	
200	mg	Thiamine-HCl x 2 H2O	
1000	ml	Distilled water