The influence of aqueous copper speciation on bioluminosity

Studies were carried out on Luria Bertani (LB) nutrient broth, typical of g rowth media used to grow strains of luminescent bacteria such as E. coil pC ulux, in order to ascertain the chemical speciation of added copper and inf er the effect of speciation on luminescent response. Both acid-base and com plexometric (with Cu) titrations were conducted. Data were analyzed using t he binding spectrum approach in which the concentration and pK value of the reactions of interest are estimated. Results of acid-base titration indica ted that LB contains both inorganic and a suite of organic functional group s, the latter probably associated with proteins and amino acids contained i n the materials used in making the medium. Titration with cupric ion sugges ted the presence of two copper-binding groups for freshly prepared LB, but three groups for media in which the bacteria had been allowed to grow for s everal hours, indicating either the introduction of a Cu-binding substance associated with microbial metabolism, or the ability to detect a weaker bin ding group as the concentrations of the other groups declined during degrad ation. Organic-Cu complexes dominate the computed speciation of Cu in these systems. Interpretation of luminescence in such systems is complicated by temporal changes in ligand type and concentration, and differential complex ation of Cu by differing ligands. If free cupric ion is assumed to be the a ctivating form for luminescence, these results indicate that the bacteria a re approximately six orders more sensitive in detecting Cu than results bas ed on total Cu alone.