In situ fluorescence cell mass measurements of Saccharomyces cerevisiae using cellular tryptophan.

This work describes a new spectroscopic optical fiber/rod technique for in situ real time measurement of cell mass and product concentrations in bioreactors using intrinsic fluorescence. The variable excitation/emission wavelength capability of this sensor allows for species-selective measurement during fermentations. Cell mass (tryptophan) and product concentrations (pyridoxine) have been measured during fermentations of Saccharomyces cerevisiae. The effects of varying substrate concentration and oxygen concentration on the observed cell mass signals are eliminated by direct measurement of cell mass, as opposed to indirect measurement schemes such as those using NADH fluorescence. The sensor is robust and able to undergo many cycles of in situ steam sterilization without degradation, and its fluorescence signal is linear with concentration for all species studied in this work. Tryptophan fluorescence from yeast is shown to be a better measure of cell mass than NADH fluorescence.

Nephrotoxicity of the 1:1 acrolein-glutathione adduct in the rat.

Previous metabolic studies in rats have suggested in vivo formation of the acrolein-glutathione (acrolein-GSH) adduct following administration of the highly reactive alpha, beta-unsaturated aldehyde acrolein. Early studies by several investigators demonstrated that similar compounds such as alpha, beta-unsaturated aldehyde-cysteine adducts have toxic (carcinostatic) activity against Ehrlich ascites tumor cells implanted in mice. The current studies investigated the in vivo toxicity associated with the acrolein-GSH adduct in the male Sprague-Dawley rat. The 1:1 acrolein-GSH adduct was synthesized and characterized by physical-chemical methods. Rats given the acrolein-GSH adduct intravenously at 0.5 or 1 mmol/kg developed nephrotoxicity characterized by glucosuria, proteinuria, elevation in serum urea nitrogen, and gross and histologic changes of the kidney. The toxicity was not affected by pretreatment of rats with pyrazole, an alcohol dehydrogenase inhibitor; disulfiram, an inhibitor of aldehyde dehydrogenases; or probenecid, a renal organic anion transport inhibitor. Administration of a similar but nonaldehydic glutathione conjugate, S-n-propylglutathione, did not result in nephrotoxicity in the rat. The nephrotoxicity induced by the acrolein-GSH adduct was inhibited by acivicin, a gamma-glutamyl-transpeptidase inhibitor. These results indicate that the acrolein-GSH adduct requires processing through the first step of the renal mercapturic acid synthesis pathway to be activated to a toxic species.

Monitoring cell concentration and activity by multiple excitation fluorometry.

Four key cellular metabolic fluorophores--tryptophan, pyridoxine, NAD(P)H, and riboflavin--were monitored on-line by a multiple excitation fluorometric system (MEFS) and a modified SLM 8000C scanning spectrofluorometer in three model yeast fermentation systems--bakers' yeast growing on glucose, Candida utilis growing on ethanol, and Saccharomyces cerevisiae RTY110/pRB58 growing on glucose. The measured fluorescence signals were compared with cell concentration, protein concentration, and cellular activity. The results indicate that the behavior and fluorescence intensity of various fluorophores differ in the various fermentation systems. Tryptophan fluorescence is the best signal for the monitoring of cell concentration in bakers' yeast and C. utilis fermentations. Pyridoxine fluoresce is the best signal for the monitoring of cell concentration in the S. cerevisiae RTY110/pRB58 fermentation. In bakers' yeast fermentations the pyridoxine fluorescence signal can be used to monitor cellular activity. The NAD(P)H fluorescence signal is a good indicator of cellular activity in the C. utilis fermentation. For this fermentation NAD(P)H fluorescence can be used to control ethanol feeding in a fed-batch process.

Temperature measurements in flames using thermally assisted laser-induced fluorescence of Ga.

The use of thermally assisted fluorescence (THAF) for temperature measurements has been investigated in a laminar, premixed C(2)H(2)/O(2)/Ar flame seeded with Ga atoms. Average temperature measurements were made with an uncertainty of less than +/-100 K in flames >2150 K and were found to be in agreement with sodium line reversal temperature measurements and equilibrium calculations. In both fuel rich and lean flames spanning equivalence ratios from 0.75 to 2.0, it was found that composition influenced the measured temperatures, resulting in an accuracy of +/-100 K over this range of flame conditions. Dilution of the flame with N(2) rather than Ar resulted in measured temperatures that were substantially higher than the calculated adiabatic flame temperature, indicating that, in this case, a partial Boltzmann equilibrium is not established among the collisionally populated levels of Ga used for the measurement. These results indicate that THAF with gallium as the thermometric species is limited to cases in which an inefficient quencher, such as a rare gas, is the primary diluent.

Remote temperature measurements in gas and gas-coal flames using the OH(0,0) middle-UV band.

This paper proposes and explores a technique for measuring the temperature of flames based on a ratio measurement of middle-ultraviolet emissions of OH. This method requires only a moderate-resolution mono-chromator, which should make it useful in remote sensing with a mobile facility. Initial experiments indicate the validity of this ratio method even when the flame has high visible luminosity due to the burning of pulverized coal. An empirical formula relates the theoretical ratio of two peak emissions to the temperature with good accuracy.