Membrane Introduction mass spectrometry with flow Injection analysis sampling has been utilized for on-line monitoring of the major products and the volatile metabolites of fermentation by the Bacillus polymyxa and Klebsiella oxytoca organisms. A flow Injection sampling system was used to rapidly deliver fermentation broth or an external standard to the mass spectrometer. Analyte Introduction occurred via a direct Insertion membrane probe In which the aqueous solutions flowed past a membrane located within the Ion source of the mass spectrometer. For both organisms, concentrations of the liquid-phase products, acetic acid, acetoln, 2,3-butanediol, and ethanol, were monitored as a function of time after permeation through the membrane and ionization by chemical Ionization. Tandem mass spectrometry confirmed that these measurements were made without Interference. Off-line gas chromatography was utilized to test the accuracy of these measurements, and excellent agreement was found. Dissolved oxygen In the fermentation broth was also monitored, and carbon dioxide and oxygen were followed In the offgases. The 02 and C02 measurements were compared with other common measurement techniques (galvanic 02 electrode, Infrared absorption, and paramagnetic 02 analysis), and very good agreement was found. The use of tandem mass spectrometry has allowed the detection of additional compounds that were previously not known to be present In measurable amounts.
Organic compounds present in aqueous solutions can be analyzed directly using a quadruple ion trap detector (ITD) when the solution is introduced via a hollow fiber membrane probe. The flow-through configuration used for sample introduction allows the aqueous solution to flow through the capillary membrane tubing while the organic compounds which selectively permeate the membrane are ionized in the ion source. In this mode of operation, the instrument shows high sensitivity. Chemical ionization mass spectra for a set of ten organic compounds of environmental interest were recorded and the ITD/membrane system was found to consistently allow detection of part-per-billion (ppb) levels of tbese compounds directly from water without any preconcentration. Analysis of well water samples containing ppb to part-per-million levels of organics was demonstrated using the ITD/membrane system. The combination of the membrane probe and ion trap produces a compact, inexpensive, rapid and sensitive system for environmental analysis. The flow-through membrane configuration was also used with a direct insertion probe in a triple quadrupole. Detection limits in the ppb range for organic compounds in water were measured. Detection of particular compounds in complex matrices was demonstrated by detection of 10 ppb 2-methoxypyridine in a fermentation medium using a triple-quadrupole mass spectrometer.
Membrane introduction mass spectrometry has been employed for on-line determination of the major products and volatile metabolitas of Bacillus polymyxa fermentation. Samples were introduced into the mass spectrometer via a direct insertion membrane probe in which the aqueous solution flowed p=u;t a membrane Iocltad in the ion source of the mass spectrometer. Concentrations of the products 2,3-butanediol, acatoin, ethanol and acetic acid in fermentation broth were measured by tandem mass spectrometry after permeation through the membrane and ionization by chemical ionization. Extern=d standards were employed for qu-,ntJflcation and = large line~ response range was available for each of the major products observed. Dissolved CO 2 and O 2, -,, well as CO 2 in the off gases, were ,,luo monitored on-line by mm spectrometry. The use of tandem mass spect:ometly has =llowod the identification of products that were not previously known to be present in measurable amounts.
Human serum butyrylcholinesterase (EC 3.1.1.8) loses 100% of its activity toward butyrylthiocholine in 60 min at pH 3.0. This deactivation is retarded by 1.37 M ammonium sulfate to a loss of 40% after 60 min at pH 3.0. Reneutralization experiments suggest that the mechanism for this acid inactivation does not exclusively involve hydrolysis of peptide bonds or protonation of the enzyme's active site. Studies with different anions and cations demonstrate that the order of their effectiveness as protective agents against acid inactivation closely follows the Hofmeister series. No relationship was found between catalytic activation or inhibition by salt and protection from acid inactivation. Ultraviolet difference studies at 288 nm with enzyme brought to pH 2.7 from pH 8.0 in the presence and absence of 1.37 M ammonium sulfate demonstrated no change in UV absorbance with ammonium sulfate present and approximately a 0.15 ODU rise in absorbance in the absence of ammonium sulfate. These results suggest that acidic pH conditions result in deactivating stereochemical changes in the active site of butyrylcholinesterase and that certain anions and cations, according to the Hofmeister series, are able to protect the enzyme from acid inactivation by stabilizing the active conformation of its active site.
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