Evidence from liposome encapsulation for transport-limited microbial metabolism of solid alkanes.

The recalcitrance of xenobiotics may be caused by an absence of transforming enzymes or by their inability to enter microbial cells. A nondestructive method for differentiating between these two possibilities is described. The solid n-alkanes octadecane (C(18)) and hexatriacontane (C(36)) were encapsulated into phosphatidylcholine bilayers (liposomes). The uptake and metabolism rates of encapsulated and unencapsulated substrates were then compared. During 1 h at 25 degrees C, a Pseudomonas isolate took up 1.3% of radiolabeled and unencapsulated C(18) (solid state) versus 23.5% of labeled and encapsulated C(18). Growth at 25 degrees C occurred with an apparent k(s) of 2453 +/- 148 mg/liter. Liposome encapsulation decreased this K(s) to 60 +/- 12 mg/liter. At 34 degrees C, growth on C(18) (liquid state) occurred with an apparent K(s) of 819 +/- 83 mg/liter and on the readily available carbon source succinate, K(s) values were 80 +/- 10 and 13 +/- 7 mg/liter at 25 and 34 degrees C, respectively. At 25 degrees C, the isolate grew on C(36) with an apparent K(s) of 2,698 +/- 831 mg/liter. Liposome encapsulation decreased the K(s) more than 60-fold to 41 +/- 7 mg/liter, resulting in the complete utilization of 400 mg of C(36) per liter in 16 h. Since controls excluded the metabolic utilization of phosphatidylcholine, the results clearly identify transport limitation as the cause for C(36) recalcitrance.