# 1290: ALKALIPHILUS MEDIUM


## Main sol. 1290

950	ml	**Solution A**	
25	ml	**Solution B**	
1	ml	**Solution C**	
25	ml	**Solution D**	

1. Dissolve ingredients of solution A, adjust pH to 8.0 with 1 N NaOH and boil medium for 1 min, then cool to room temperature under 100% N2 gas atmosphere. Dispense solution A under same gas atmosphere in anoxic culture vessels (e.g., Hungate-type tubes) and autoclave. Prior to inoculation complete the medium by adding appropriate amounts of solutions B, C and D prepared under 100% N2 gas and sterilized by filtration. Adjust pH of the completed medium to 8.0 with a sterile anoxic solution of 0.1 N HCl or 0.1 N NaOH, if necessary.


## Solution A

75	g	NaCl	
8.8	g	MgSO4 x 7 H2O	
1	g	KCl	
10	g	Yeast extract (OXOID)	
10	g	Trypticase peptone (BD BBL)	
10	ml	**Modified Wolin's mineral solution**	
940	ml	Distilled water	


## Solution B

2.5	g	D-Glucose	
25	ml	Distilled water	


## Solution C

1	ml	**Wolin's vitamin solution (10x)**	


## Solution D

0.5	g	Na-thioglycolate	
0.5	g	Na-ascorbate	
25	ml	Distilled water	


## Modified Wolin's mineral solution

1.5	g	Nitrilotriacetic acid	
3	g	MgSO4 x 7 H2O	
0.5	g	MnSO4 x H2O	
1	g	NaCl	
0.1	g	FeSO4 x 7 H2O	
0.18	g	CoSO4 x 7 H2O	
0.1	g	CaCl2 x 2 H2O	
0.18	g	ZnSO4 x 7 H2O	
0.01	g	CuSO4 x 5 H2O	
0.02	g	AlK(SO4)2 x 12 H2O	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.03	g	NiCl2 x 6 H2O	
0.3	mg	Na2SeO3 x 5 H2O	
0.4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water	

1. First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.


## 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