Biodiesel Glycerol Byproduct as a Feedstock for Pseudomonads of Industrial Significance in Undergraduate Research

Graves, James, Christopher Fronczak, Kezia Mathew, and Seon Gyeong Park

In manufacture of biodiesel a byproduct is made that contains glycerol.  Pseudomonas species are used to synthesize biochemicals and in bioremediation.  Because some isolates of Pseudomonas utilize glycerol the growth of selected species with biodiesel glycerol byproduct was investigated.  When assessed by a rapid solid phase lysochrome diazo dye assay (Precision Labs, Inc.), glycerol, byproduct made by transesterification or catalyst column, and soap failed to test positive for lipid in contrast to vegetable oil, biodiesel and tween 20.  On spirit blue agar Pseudomonas species exhibited a halo around areas of bacterial growth which was indicative of the presence of lipase.  Ability to grow at 14oC or 42oC differentiated several species.  Antibiograms made by disk diffusion showed all the bacteria exhibited sensitivity to ciprofloxacin, gentamicin and tetracycline, but some grew up to disks of chloramphenicol and erythromycin.  On minimal agar, containing only essential nutrients, with glucose, glycerol or byproduct made by transesterification or catalyst column as carbon sources, P. aeruginosa, P. cepacia, P. fluorescens and P. putida showed fair to excellent growth.  In minimal broth, when glucose, glycerol or byproduct from catalyst column served as carbon source, measurement of optical density (OD) by a Klett – Summerson colorimeter showed P. aeruginosa, P. fluorescens and P. putida attained ODs over 150. With byproduct made by transesterification, P. aeruginosa, and P. putida produced ODs near 100 and P. fluorescens showed an OD near 50.  Viable cell counts in medium with byproduct attained 108 colony forming units (CFU)/ml.  When tested in a microplate redox-based array (Biolog, Inc.) the bacteria demonstrated ability for oxidation of glycerol and tweens.  Biodiesel glycerol byproduct could serve as a possible carbon source for Pseudomonas species.