# 386: DESULFOLUTIVIBRIO MEDIUM


## Main sol. 386

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

1. Solution A is sparged 80% N2 and 20% CO2 gas mixture for 30 – 45 min to make it anoxic, then distributed under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclaved. Solution B is autoclaved separately under 80% N2 and 20% CO2 gas mixture. Solutions C and F are autoclaved under 100% N2 gas. 
Solutions D and E are prepared under 100% N2 gas atmosphere and sterilized by filtration. To complete the medium appropriate amounts of the solutions B to F are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be at 7.1 - 7.4.
2. Note: When growth has started feed culture again with same amount of pyrosulfite. After a further two days repeat feeding once more.


## Solution A

0.2	g	KH2PO4	
0.25	g	NH4Cl	
1	g	NaCl	
0.4	g	MgCl2 x 6 H2O	
0.5	g	KCl	
0.15	g	CaCl2 x 2 H2O	
1	ml	**Trace element solution SL-10**	
1	ml	**Selenite-tungstate solution**	
0.5	ml	Sodium resazurin (0.1% w/v)	
940	ml	Distilled water	


## Solution B

1.5	g	Na2CO3	
30	ml	Distilled water	


## Solution C

0.3	g	Na-acetate x 3 H2O	
10	ml	Distilled water	


## Solution D

1	ml	**Seven vitamins solution**	


## Solution E

0.95	g	Na2S2O5 (sodium pyrosulfite)	
10	ml	Distilled water	

1. Adjust pH to 7.5-8.0 with NaOH.


## Solution F

0.4	g	Na2S x 9 H2O	
13	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.


## Selenite-tungstate solution

0.5	g	NaOH	
3	mg	Na2SeO3 x 5 H2O	
4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water	


## 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