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Feed medium also contained trace elements and vitamins (concentrated four-fold compared to medium for batch cultures) and Clerol FBA 3107 antifoam (1 mL L−1).

The medium for batch and continuous cultivation was Minimal Medium and contained per liter: 4.5 g NH4Cl, 1.5 g KH2PO4, 0.5 KCl, 0.5 MgSO4·7H2O and 1 ml trace metal solution.

The medium for batch cultures in shake flasks contained (g L-1): glycerol 10, NZ-amine 5, (NH4 2HPO4 4, KH2PO4 5, K2HPO4 7.4, MgSO4 * 7H2O 1.2, thiamine HCl 0.015, ampicillin 0.1 and 10 mL of trace element solution (TES).

The medium for batch cultures in the bioreactor contained (g L-1): glycerol 20, NZ-Amine 5, (NH4 2HPO4 4, KH2PO4 13.3, (NH4 2SO4 1, MgSO4 * 7H2O 1.2, thiamine HCl 0.015, ampicillin 0.1 and 10 ml of TES containing additional 0.15 g l-1 CuCl2 * 4H2O.

Pre-inoculum was made starting from one vial of the Molecular Biology cell bank in culture medium FMYb, an animal free derived media composed by a pre defined solution of yeast extract, glycerol, vitamins and minerals with apposite antibiotic and then inoculated in 10 litres of FMYb culture medium for batch fermentation.

The standard production medium for batch cultures contained the following ingredients in 1 L of distilled water (modified from Biebl 2001): glycerol, varied; glucose, varied; K2HPO4, 0.5 g; KH2PO4, 0.5 g, MgSO4 · 7H20, 0.2 g; (NH4 2SO4, 5 g; CaCl2 2H2O, 0.02 g, FeSO4 7H2O, 0.01 g; cysteine HCl, 0.3 g; resarzurin, 0.005 g; 2 mL of trace element solution SL7 and 1 g yeast extract.

Similar(54)

Unless otherwise indicated, all media used was the inorganic nitrogen-limiting (0.15 g L-1 (NH4)2SO4) medium used for batch and chemostat cultures described by Verduyn et al. [ 69].

Cultures were grown in the same MTC medium used for batch studies, with 3 g of switchgrass and with reducing agents and vitamins added separately after the autoclaving of fermentor vessels.

The feed-medium had the same composition as the medium for the batch phase, except as carbon source a mixture of 20 g L-1 glucose and 50 g L-1 xylose were used.

The medium for the batch cultures contained molasses (245 g/L, sucrose content of 600 g/L, Lesaffre, Rangueil, France), NH4Cl (4.0 g/L), KH2PO4 (0.5 g/L), MgCl2 (1.0 g/L), and YNB (without amino acids and ammonium sulfate, 1.5 g/L, Difco).

The cell pellets and the immobilized cells were collected at the 50 micron nylon mesh screen at the sampling port and replenished with fresh fermentation medium for further batches.

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