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Dissolve ingredients (except ferric citrate, carbonate and vitamins) and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add ferric citrate (dissolve by boiling and adjust pH to 6.5 - 7.0) and vitamins from sterile anoxic stock solutions prepared under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Sterilize vitamins by filtration. Adjust pH of the complete medium to 6.5, if necessary.
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