Kirit D. Chapatwala

Biodegradation of cyanides, cyanates and thiocyanates to ammonia and carbon dioxide by immobilized cells of Pseudomonas putida

Pseudomonas putida utilizes cyanide as the sole source of carbon and nitrogen. Agar, alginate, and carrageenan were screened as the encapsulating matrices for P. putida. Alginate-immobilized cells of P. putida degraded sodium cyanide (NaCN) more efficiently than non-immobilized cells or cells immobilized in agar or carrageenan. The end products of biodegradation of cyanide were identified as ammonia (NH3) and carbon dioxide (CO2). These products changed the medium pH. In bioreactors, the rate of cyanide degradation increased with an increase in the rate of aeration. Maximum utilization of cyanide was observed at 200 ml min−1 of aeration. Immobilized cells of P. putida degraded cyanides, cyanates and thiocyanates to NH3 and CO2. Use of Na[14C]-CN showed that 70% of carbon of Na[14C]-CN was converted into 14CO2 and only 10% was associated with the cell biomass. The substrate-dependent kinetics indicated that the Km and Vmax values of P. putida for the substrate, NaCN were 14 mM and 29 nmol of oxygen consumed mg protein−1 min−1 respectively.

Pseudomonas marginalis: its degradative capability on organic nitriles and amides.

Pseudomonas marginalis, capable of utilizing acetonitrile as the sole source of carbon and nitrogen, was isolated from an industrial waste site. P. marginalis metabolized acetonitrile into ammonia and acetate. The minimal inhibitory concentration values of different nitriles and amides for P. marginalis were in the range 5–300 mM. The bacterium was able to transform high-molecular-mass nitrile compounds and their respective amides into ammonia. The data from substrate-dependent kinetics showed that the Km and Vmax values of P. marginalis for acetonitrile were 33 mM and 67 nmol oxygen consumed min−1 (ml cell suspension)−1 respectively. The study with [14C]acetonitrile indicated that nearly 66% of the carbon was released as 14CO2 and 12% was associated with the biomass. The enzyme system involved in the hydrolysis of acetonitrile was shown to be intracellular and inducible. The specific activities of the enzymes nitrile aminohydrolase and amidase were determined in the cell-free extracts of P. marginalis. Both the enzymes could hydrolyze a wide range of nitriles and amides. The present study suggests that the biodegradation of organic nitriles and the bioproduction of organic acids may be achieved with the cells of P. marginalis.

Degradation of organic cyanides by pseudomonas aeruginosa

A bacterium capable of utilizing acetonitrile (methyl cyanide) as the sole source of carbon and nitrogen was isolated from soil and identified asPseudomonas aeruginosa. This bacterium could also utilize and oxidize numerous lower-mol-wt nitrile compounds and their corresponding amides as growth substrates. A metabolite of acetonitrile in the culture medium was determined to be ammonia. The accumulation of ammonia in the culture medium was proportional to the concentration of the substrate and the inoculum. Cell extracts of the bacterium contained activities corresponding to nitrile aminohydrolase (E C 3.5.5.1) and amidase (E C 3.5.1.4), which regulate the degradation of acetonitrile. Both enzymes were inducible and hydrolyzed a wide range of substrates, and it was determined that the specific activity of amidase was far greater than the activity of nitrile aminohydrolase.

Conversion of sodium cyanide to carbon dioxide and ammonia by immobilized cells ofPseudomonas putida

SummaryPseudomonas putida, isolated from contaminated industrial wastewaters and soil sites, was found to utilize sodium cyanide (NaCN) as a sole source of carbon and nitrogen. Cells, immobilized in calcium alginate beads (1–2 mm diameter) were aerated in air-uplift-type fluidized batch bioreactor containing 100–400 ppm of NaCN. Degradation of NaCN was monitored for 168 h by analyzing gaseous and dissolved ammonia (NH3), CO2, pH and optical density. The results indicated that the alginate-immobilized cells ofP. putida were able to degrade NaCN into NH3 and CO2 in a time-dependent manner.

Cell-free extract(s) of Pseudomonas putida catalyzes the conversion of cyanides, cyanates, thiocyanates, formamide, and cyanide-containing mine waters into ammonia

Our isolate, Pseudomonas putida, is known to be capable of utilizing cyanides as the sole source of carbon (C) and nitrogen (N) both in the form of free cells and cells immobilized in calcium alginate. In the present study, the cell-free extract(s) were prepared from the cells of P. putida grown in the presence of sodium cyanide. The ability of enzyme(s) to convert cyanides, cyanates, thiocyanates, formamide and cyanide-containing mine waters into ammonia (NH3) was studied at pH 7.5 and pH 9.5. The kinetic analysis of cyanide and formamide conversion into NH3 at pH 7.5 and pH 9.5 by the cell-free extract(s) of P. putida was also studied. The Km and Vmax values for cyanide/formamide were found to be 4.3/8 mM and 142/227 μmol NH3 released mg protein−1 min−1 respectively at pH 7.5 and 5/16.67 mM and 181/434 μmol NH3 released mg protein−1 h−1 respectively at pH 9.5. The study thus concludes that the cell-free extract(s) of P. putida is able to metabolize not only cyanides, cyanates, thiocyanates, and formamide but also cyanide-containing mine waters to NH3.

Effect of cadmium on hepatic and renal gluconeogenic enzymes in female rats

Earlier, we have reported that cadmium (Cd) induced gluconeogenesis in male rats. Since females are as much exposed to cadmium as are males, this study was conducted to determine Cd effects on gluconeogenesis in female rats. Adult female rats were injected intraperitoneally (i.p.) with Cd at dose levels of 0.25, 0.75 and 1.25 mg/kg body weight per day for 4 weeks. The controls received saline for the same length of time. Daily food consumption and body weight gain were recorded. At the end of 2 and 4 weeks, 4 rats from each group were killed. Extension of treatment with 1.25 mg Cd for 4 weeks resulted in extreme Cd toxicity killing all animals before the completion of full treatment period. There were no significant changes in total body weight gain and weights of liver and kidney due to Cd. Serum protein increased significantly in animals receiving 0.75 and 1.25 mg Cd for 4 and 2 weeks, whereas serum glucose increased only in animals injected with 1.25 mg Cd for 2 weeks. SGOT and SGPT were elevated (P less than 0.01) in dose- and time-dependent fashion. Activities of three key gluconeogenic enzymes glucose-6-phosphatase (G-6-Pase), fructose-1,6-diphosphatase (FD-Pase), and phosphoenolpyruvate carboxykinase (PEPCK) in liver and kidney were induced significantly (P less than 0.01) in animals injected with 0.75 mg for 2 and 4 weeks and 1.25 mg for 2 weeks, and these increases were dose- and time-related. These results suggest that Cd alters hepatic and renal gluconeogenesis in female rats also.

Screening of encapsulated microbial cells for the degradation of inorganic cyanides

SummaryDifferent encapsulation matrices were screened to encapsulate cells ofPseudomonas putida for degradation of inorganic cyanides. Degradation of NaCN by free cells and cells immobilized in agar, alginate or carrageenan matrices was studied. The rate of NaCN degradation was monitored for 120 h by measuring pH, bacterial growth, dissolved and gaseous NH3 and gaseous CO2. Alginate-immobilized cells degraded NaCN more efficiently than free cells or agar- or carrageenan-immobilized cells.

Isolation and characterization of acetonitrile utilizing bacteria.

SummaryBacteria utilizing high concentrations of acetonitrile as the sole carbon source were isolated and identified asChromobacterium sp. andPseudomonas aeruginosa. Maximum growth was attained after 96 h of incubation andP. aeruginosa grew slightly faster thanChromobacterium sp. The strains were able to grow and oxidize acetonitrile at concentrations as high as 600 mM. However, higher concentrations inhibited growth and oxygen uptake. Degradation studies with (14C)acetonitrile indicated 57% of acetonitrile was degraded byPseudomonas aeruginosa as compared to 43% byChromobacterium. The isolates utilized different nitrile compounds as carbon substrates.

A kinetic study on the bioremediation of sodium cyanide and acetonitrile by free and immobilized cells of Pseudomonas putida

Pseudomonas putida capable of utilizing organic nitrile (acetonitrile) and inorganic cyanide (sodium cyanide) as the sole source of carbon and nitrogen was isolated from contaminated industrial sites and waste water. The bacterium possesses nitrile aminohydrolase (EC 3.5.5.1) and amidase (EC 3.5.1.4), which are involved in the transformation of cyanides and nitriles into ammonia and CO2 through the formation of amide as an intermediate. Both of the enzymes have a high selectivity and affinity toward the−Cn group. The rate of degradation of aceotnitrile and sodium cyanide to ammonia and CO2 by the calcium-alginate immobilized cells ofP. putida was studied. The rate of reaction during the biodegradation of acetonitrile and sodium cyanide, and the substrate- and product-dependent kinetics of these toxic compounds were studied using free and immobilized cells ofP. putida and modeled using a simple Michaelis-Menten equation.

Degradative capability of Pseudomonas putida on acetonitrile.

Pseudomonas putida, capable of utilizing acetonitrile as a sole source of carbon and nitrogen, was isolated from contaminated soil and water samples collected from industrial sites. TheP. putida cells were immobilized in calcium alginate beads. The degradation of acetonitrile by the immobilized cells ofP. putida was investigated. The immobilized cells degraded different concentrations of acetonitrile into ammonia and carbon dioxide. The effect of aeration on the degradation rate was also studied. Oxygen limitation was suggested in the alginate-immobilized system. The rate of degradation of acetonitrile increased with increase in the rate of aeration.