# 845: SPIRILLUM WINOGRADSKYI MEDIUM


## Main sol. 845

5	g	Peptone (BD Bacto)	
1	g	Disodium succinate	
1	g	(NH4)2SO4	
1	g	MgSO4 x 7 H2O	
2	mg	FeCl3 x 6 H2O	
2	mg	MnSO4 x H2O	
1	g	Agar (BD Bacto)	
1	ml	**Trace element solution SL-11**	
1	ml	**Seven vitamins solution**	
1	ml	Catalase (2000 U/ml)	
1000	ml	Distilled water	

1. Dissolve ingredients except trace elements, vitamins and catalase, adjust pH to 7.2 - 7.4, bring medium to the boil and cool to room temperature under 100% N2 gas atmosphere. Dispense under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. To complete the medium add trace elements, vitamins and catalase from sterile anoxic stock solutions prepared under 100% N2 gas atmosphere. Vitamins and catalase should be sterilized by filtration. After inoculation, inject sterile air into the culture vessel up to about 20% of the total gas volume (i.e., 4 parts N2 and 1 part air).


## Trace element solution SL-11

5.2	g	Na2-EDTA x 2 H2O	
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	
1000	ml	Distilled water	

1. Dissolve EDTA in 800 ml distilled water, adjust pH to 7 using 2 N NaOH and add ferrous chloride. After ferrous chloride has dissolved add other compounds. Finally adjust pH to 6.0 and bring volume to 1000 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