Characterization of Enterobacter cloacae BAGM01 Producing a Thermostable and Alkaline-Tolerant Rhamnolipid Biosurfactant from the Gulf of Mexico.

The search for novel biosurfactants (Bs) requires the isolation of microorganisms from different environments. The Gulf of Mexico (GoM) is a geographical area active in the exploration and exploitation of hydrocarbons. Recent metagenomic and microbiologic studies in this area suggested a potential richness for novel Bs microbial producers. In this work, nineteen bacterial consortia from the GoM were isolated at different depths of the water column and marine sediments. Bs production from four bacterial consortia was detected by the CTAB test and their capacity to reduce surface tension (ST), emulsion index (EI24), and hemolytic activity. These bacterial consortia produced Bs in media supplemented with kerosene, diesel, or sucrose. Cultivable bacteria from these consortia were isolated and identified by bacterial polyphasic characterization. In some consortia, Enterobacter cloacae was the predominant specie. E. cloacae BAGM01 presented Bs activity in minimal medium and was selected to improve its Bs production using a Taguchi and Box-Behnken experimental design; this strain was able to grow and presented Bs activity at 35 g L-1 of NaCl. This Bs decreased ST to around 34.5 ± 0.56 mNm-1 and presented an EI24 of 71 ± 1.27%. Other properties of this Bs were thermal stability, stability in alkaline conditions, and stability at high salinity, conferring important and desirable characteristics in multiple industries. The analysis of the genome of E. cloacae BAGM01 showed the presence of rhlAB genes that have been reported in the synthesis of rhamnolipids, and alkAB genes that are related to the degradation of alkanes. The bioactive molecule was identified as a rhamnolipid after HPLC derivatization, 1H NMR, and UPLC-QTOF-MS analysis.

Microbial prospection of communities that produce biosurfactants from the water column and sediments of the Gulf of Mexico.

Microbial communities capable of hydrocarbon degradation linked to biosurfactant (BS) and bioemulsifier (BE) production are basically unexplored in the Gulf of México (GOM). In this work, the BS and BE production of culturable marine bacterial hydrocarbonoclasts consortia isolated from two sites (the Perdido Fold Belt and Coatzacoalcos area) was investigated. The prospection at different locations and depths led to the screening and isolation of a wide variety of bacterial consortia with BS and BE activities, after culture enrichment with crude oil and glycerol as the carbon sources. At least 55 isolated consortia presented reduction in surface tension (ST) and emulsifying activity (EI24 ). After colony purification, bacteria were submitted to polyphasic analysis assays that resulted in the identification of different strains of cultivable Gammaproteobacteria Gram (-) Citrobacter, Enterobacter, Erwinia, Pseudomonas, Vibrio, Shewanella, Thalassospira, Idiomarina, Pseudoalteromonas, Photobacterium, and Gram (+) Staphylococcus, Bacillus, and Microbacterium. Overall, the best results for ST reduction and EI24 were obtained with consortia. Individually, Pseudomonas, Bacillus, and Enterobacter strains showed the best results for the reduction of ST after 6 days, while Thalassospira and Idiomarina strains showed the best results for EI24 (above 68% after 9 days). Consortia isolates from the GOM had the ability to degrade crude oil by up to 40-80% after 24 and 36 months, respectively. In all cases, biodegradation of crude oil was related to the reduction in ST and bioemulsifying activity and was independent from the depth in the water column.

Improved production, purification, and characterization of biosurfactants produced by Serratia marcescens SM3 and its isogenic SMRG-5 strain.

In this study, the biosurfactants (Bs) production of two Serratia marcescens strains (SM3 and its isogenic SMRG-5 strain) was improved and the tenso-active agents were purified and characterized. A 23 factorial design was used to evaluate the effect of nitrogen and carbon sources on the surface tension (ST) reduction and emulsion index (EI24 ) of the produced Bs. Optimum Bs production by SM3 was achieved at high concentrations of carbon and nitrogen, reducing ST to 26.5 ± 0.28 dynes/cm, with an EI24 of 79.9 ± 0.2%. Meanwhile, the best results for SMRG-5 were obtained at low concentrations, reducing the ST to 25.2 ± 0.2 dynes/cm, with an EI24 of 89.7 ± 0.28%. The optimal conditions for Bs production were scaled up in a 2-L reactor, yielding 4.8 and 5.2 g/L for SM3 and SMRG-5, respectively. Gas Chromatography-Mass Spectrometry (GC-MS) analysis revealed the presence of two different lipopeptides (hidrofobic fractions: octadecanoic and hexadecanoic acid for SM3 and SMRG5, respectively). Both strains were capable of benzo [a] pyrene removal (59% after 72 H of culture).

Production and application of a thermostable lipase from Serratia marcescens in detergent formulation and biodiesel production.

In this study, extracellular lipase was produced by Serratia marcescens wild type and three mutant strains. The maximum lipase activity (80 U/mL) was obtained with the SMRG4 mutant strain using soybean oil. Using a 22 factorial design, the lipase production increased 1.55-fold (124 U/mL) with 4% and 0.05% of soybean oil and Triton X-100, respectively. The optimum conditions for maximum lipase activity were 50 °C and pH 8. However, the enzyme was active in a broad range of pH (6-10) and temperatures (5-55 °C). This lipase was stable in organic solvents and in the presence of oxidizing agents. The enzyme also proved to be efficient for the removal of triacylglycerol from olive oil in cotton cloth. A Box-Behnken experimental design was used to evaluate the effects of the interactions between total lipase activity, buffer pH, and wash temperatures on oil removal. The model obtained suggested that all selected factors had a significant impact on oil removal, with optimum conditions of 550 U lipase, 45 °C, pH 9.5, with 79.45% removal. Biotransformation of waste frying oil using the enzyme and in presence of methanol resulted in the synthesis of methyl esters such as methyl oleate, methyl palmitate, and methyl stearate.