Dario Frascari, Davide Pinelli, Massimo Nocentini, Stefano Fedi, Youri Pii, Davide Zannoni

Chloroform degradation by butane-grown cells of Rhodococcus aetherovorans BCP1

Electronic Supplementary Material

S1 Experimental details relative to the resting cells assays

Vial volume / 11.6 ml in all assays except in n. 15 (119 ml)
Phosphate buffer volume / Assays n. 4, 5, 6, 7, 9, 10, 12, 13, 14: 1 ml
Assays n. 1, 2, 11, 16, 17, 18, 19: 4 ml
Assays n. 3, 8: 6 ml
Assays n. 15: 40 ml
Phosphate buffer composition / K2HPO4 8900 mM, NaH2PO4·H2O 5355 mM
Inoculum volume / ¼ of phosphate buffer volume
Operational procedure for preparation of the inoculum / Strain BCP1 was grown in ten 119-ml vials each containing 60 ml of sterile mineral medium (mM: (NH4)2SO4 797, MgSO4·7H2O 244, CaCl2 132, K2HPO4 8900, NaH2PO4·H2O 5355, FeSO4·7H2O 22.6, NaNO3 9000, MnCl2·4H2O 1.52, ZnSO4·7H2O 0.510, H3BO3 1.00, Na2MO4·2H2O 0.450, NiCl2·2H2O 0.144, CuCl2·2H2O 0.100, CoCl2·6H2O 0.100) buffered with phosphate and amended with 97 mmol of 0.22 mm-filtered butane (for assays n. 1-17) or n-hexane (for assays n. 18 and 19); upon completion of growth substrate consumption, the vials were centrifuged (5000 rpm, 10 min, 10°C) and the cells were washed twice with pH-7 sterile phosphate buffer and re-suspended in the same buffer
Initial cell concentration / The initial cell concentration in the resting-cell assays varied between 75 mgprotein l-1 and 300 mgprotein l-1
Experimental details relative to the introduction of the compounds in the vials / Butane, acetylene and VC were taken from a cylinder with a sterile syringe and 0.22-mm filtered prior to introduction in the vials.
Chloroform, t-DCE and 1,1,2-TCA were taken from sterile aqueous solutions (8.1, 4.4 and 0.54 mM, respectively) with a sterile 25-ml syringe.
Allylthiourea (ATU) was taken from a 100 mM solution with a sterile 5-ml or 25-ml syringe.
Specific operational procedure relative to assays n. 7 and 8 / 1) introduction of the inoculum in the vial; 2) introduction of 0.22 mm-filtered acetylene with a sterile syringe; 3) after 15 minutes of agitation at 190 rpm and 30°C, purging of the vial with 0.22 mm-filtered air to remove the acetylene; 4) introduction of butane or chloroform; 5) first headspace analysis after 15 minutes of agitation at 190 rpm and 30°C