3-BrPA (3-bromopyruvate) is an alkylating agent with anti-tumoral activity on hepatocellular carcinoma. This compound inhibits cellular ATP production owing to its action on glycolysis and oxidative phosphorylation; however, the specific metabolic steps and mechanisms of 3-BrPA action in human hepatocellular carcinomas, particularly its effects on mitochondrial energetics, are poorly understood. In the present study it was found that incubation of HepG2 cells with a low concentration of 3-BrPA for a short period (150 microM for 30 min) significantly affected both glycolysis and mitochondrial respiratory functions. The activity of mitochondrial hexokinase was not inhibited by 150 microM 3-BrPA, but this concentration caused more than 70% inhibition of GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 3-phosphoglycerate kinase activities. Additionally, 3-BrPA treatment significantly impaired lactate production by HepG2 cells, even when glucose was withdrawn from the incubation medium. Oxygen consumption of HepG2 cells supported by either pyruvate/malate or succinate was inhibited when cells were pre-incubated with 3-BrPA in glucose-free medium. On the other hand, when cells were pre-incubated in glucose-supplemented medium, oxygen consumption was affected only when succinate was used as the oxidizable substrate. An increase in oligomycin-independent respiration was observed in HepG2 cells treated with 3-BrPA only when incubated in glucose-supplemented medium, indicating that 3-BrPA induces mitochondrial proton leakage as well as blocking the electron transport system. The activity of succinate dehydrogenase was inhibited by 70% by 3-BrPA treatment. These results suggest that the combined action of 3-BrPA on succinate dehydrogenase and on glycolysis, inhibiting steps downstream of the phosphorylation of glucose, play an important role in HepG2 cell death.

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Drilling fluid has many functions, such as carry cuttings from the hole permitting their separation at the surface, cool and clean the bit, reduce friction between the drill pipe and wellbore, maintain the stability of the wellbore, and prevent the inflow of fluids from the wellbore and form a thin, low-permeable filter cake. Filter cake removal is an important step concerning both production and injection in wells, mainly concerning horizontal completion. The drilling fluids are typically comprised of starch, the most important component of the filter cake. A common approach to remove this filter cake is the use of acid solutions. However, these are non-specific reactants. A possible alternative is the use of enzymatic preparations, like amylases, that are able to hydrolyze starch. Wells usually operate in drastic conditions for enzymatic preparations, such as high temperature, high salt concentration, and high pressure. Thus, the main objective of this work was to characterize four enzymatic preparations for filter cake removal under open hole conditions. The results showed that high salt concentrations (204,000 ppm NaCl) in completion fluid decreased amylolytic activity. All enzymatic preparations were able to catalyze starch hydrolysis at all temperatures tested (30, 65, 80, and 95 degrees C). An increase of amylolytic activity was observed with the increase of pressure (100, 500 and 1,000 psi) for one commercial amylase.