Factors influencing the trans-membrane transport of n-octadecane by Pseudomonas sp. DG17.

In soil bioremediation techniques, the trans-membrane transport of hydrocarbons across the cell membrane is a new and complex point of understanding the process of hydrocarbons biodegradation. In this study, the effect of different environmental factors, including substrate concentration, bacterial inoculums, pH, salinity, substrate analogues and nutrients, on the transport of [14C]n-octadecane by Pseudomonas sp. DG17 was investigated. The results showed that cellular [14C]n-octadecane levels increased along with the increase in the substrate concentration. However, the trans-membrane transport of [14C]n-octadecane was a saturable process in the case of equal amounts of inoculum (biomass). The highest concentration of accumulated [14C]n-octadecane was 0.51 µmol mg-1 ± 0.028 µmol mg-1 after incubation for 20 min. Meanwhile, the cellular n-octadecane concentration decreased along with the biomass increase, and reached a stable level. Acidic/alkaline conditions, high salinity, and supplement of substrate analogues could inhibit the transport of [14C]n-octadecane by Pseudomonas sp. DG17, whereas nitrogen or phosphorus deficiency did not influence this transport. The results suggested that trans-membrane transport of octadecane depends on both the substrate concentration and the microorganism biomass, and extreme environmental conditions could influence the biodegradation ability of microorganisms through inhibiting the transport of extracellular octadecane.

Immobilization of Pseudomonas sp. DG17 onto sodium alginate-attapulgite-calcium carbonate.

A strain of Pseudomonas sp. DG17, capable of degrading crude oil, was immobilized in sodium alginate-attapulgite-calcium carbonate for biodegradation of crude oil contaminated soil. In this work, proportion of independent variables, the laboratory immobilization parameters, the micromorphology and internal structure of the immobilized granule, as well as the crude oil biodegradation by sodium alginate-attapulgite-calcium carbonate immobilized cells and sodium alginate-attapulgite immobilized cells were studied to build the optimal immobilization carrier and granule-forming method. The results showed that the optimal concentrations of sodium alginate-attapulgite-calcium carbonate and calcium chloride were 2.5%-3.5%, 0.5%-1%, 3%-7% and 2%-4%, respectively. Meanwhile, the optimal bath temperature, embedding cell amount, reaction time and multiplication time were 50-60 °C, 2%, 18 h and 48 h, respectively. Moreover, biodegradation was enhanced by immobilized cells with a total petroleum hydrocarbon removal ranging from 33.56% ± 3.84% to 56.82% ± 3.26% after 20 days. The SEM results indicated that adding calcium carbonate was helpful to form internal honeycomb-like pores in the immobilized granules.

Pseudosolubilized n-alkanes analysis and optimization of biosurfactants production by Pseudomonas sp. DG17.

The pseudosolubilized medium-chain-length n-alkanes during biodegradation process, and optimization of medium composition and culture conditions for rhamnolipid production by Pseudomonas sp. DG17 using Plackett-Burman design and Box-Behnken design, were examined in this study. The results showed that pseudosolubilized concentration of C14 to C20 n-alkanes was higher than that of C24 to C26. After incubation for 120 h, pseudosolubilized C16H34 increased to 2.63 ± 0.21 mg. Meanwhile, biodegradation rates of n-alkanes decreased along with the increase of carbon chain length. Carbon-14 assay suggested that nonlabeled C14H30, C16H34, and C20H42 inhibited the biodegradation of (14)C n-octadecane, and Pseudomonas sp. DG17 utilized different alkanes simultaneously. Three significant variables (substrate concentration, salinity, and C/N) that could influence rhamnolipid production were screened by Plackett-Burman design. Results of Box-Behnken design suggested that rhamnolipid concentration could be achieved at 91.24 mg L(-1) (observed value) or 87.92 mg L(-1) (predicted value) with the optimal levels of concentration, salinity, and C/N of 400 mg L(-1), 1.5 %, and 45, respectively.

Trans-membrane transport of n-octadecane by Pseudomonas sp. DG17.

The trans-membrane transport of hydrocarbons is an important and complex aspect of the process of biodegradation of hydrocarbons by microorganisms. The mechanism of transport of (14)C n-octadecane by Pseudomonas sp. DG17, an alkane-degrading bacterium, was studied by the addition of ATP inhibitors and different substrate concentrations. When the concentration of n-octadecane was higher than 4.54 µmol/L, the transport of (14)C n-octadecane was driven by a facilitated passive mechanism following the intra/extra substrate concentration gradient. However, when the cells were grown with a low concentration of the substrate, the cellular accumulation of n-octadecane, an energy-dependent process, was dramatically decreased by the presence of ATP inhibitors, and n-octadecane accumulation continually increased against its concentration gradient. Furthermore, the presence of non-labeled alkanes blocked (14)C n-octadecane transport only in the induced cells, and the trans-membrane transport of n-octadecane was specific with an apparent dissociation constant K t of 11.27 µmol/L and V max of 0.96 µmol/min/mg protein. The results indicated that the trans-membrane transport of n-octadecane by Pseudomonas sp. DG17 was related to the substrate concentration and ATP.