news Separations on the bacterial scale Various efforts have been made to extend electrophoresis to the separation of microbes, but they have met with limited success. Of the few reports available, many concentrate on single organisms; even so, the electropherograms may have multiple peaks, which complicate the interpretation. Attempts to separate different kinds of organisms have been time-consuming and of poor efficiency and resolution. In the Dec. 15 issue of Analytical Chemistry (pp 5465–5469), Daniel Armstrong and his colleagues at the University of Missouri–Rolla describe two capillary electrokinetic approaches to separating different types of bacteria efficiently. Unlike many current methods, which use lysed cells, these techniques use intact cells, which may eliminate some of the variability usually ascribed to differing growth conditions, ages of cells, and preparation procedures. In the first approach, the researchers used capillary isoelectric focusing (CIEF) to separate the common bacteria Escherichia coli K12, Pseudomonas putida, and Serratia rubidae, all of which are rodshaped and approximately 1 µm in diameter. Samples were injected into a capillary under pressure, followed by an ampholyte to generate a pH gradient. A “focusing” voltage was applied for 5 min, and the samples were mobilized with a low-pressure rinse. Detection was at 280 nm using a standard on-line UV detector. Under these conditions, the amphoteric bacteria separated rapidly, migrating to a position
where they were uncharged. “This position can depend on the surface features characteristic of an individual species,” explains Armstrong. The second approach relied on carrying the negatively charged bacteria in a semidilute polymer solution—in this case, poly(ethylene oxide)—during CE. This method was slower but could Electropherogram showing the capillary isoelectric separate bacteria not amenable focusing separation of three bacteria of similar size. to the CIEF approach. The polymer solution was added to the running buffer for the column. The ticular, are easily damaged by oxygen, bacteria samples were injected for 8–10 s, pH, secretions from other bacteria, and and the separation was performed at 10 rough handling. In addition, a number of kV, with on-line detection at 214 nm. other factors—including the formation of The researchers tested the polymer aggregates, attachment to surfaces, and technique with a mixture of yeast sampling problems due to the relatively (Saccharomyces cerevisiae) and bacteria low concentrations of microbes in soluboth spherical (Micrococcus luteus) and tion—must be controlled to obtain reprorod-shaped (P. fluorescens and Enterducible separation results. obacter aerogenes). The microbes Nevertheless, Armstrong thinks bacappeared to separate according to size, teria are coming into analytical line with shape, and other factors, and varying the macromolecules, and he expects that concentration of the polymer changed separating microbes one day will be as the elution orders. “The exact mechaeasy as separating molecules is now. This nism of this action is not yet clear,” says kind of analysis will “revolutionize Armstrong. “Importantly, though, very aspects of microbiology involving the high separation efficiencies were obdiagnosis and profiling of some diseases, tained for both methodologies.” QC in fermentation, soil analysis, bioArmstrong points out that although remediation studies, and virtually any they have obtained decent results, the other area of science and technology process of separating microorganisms is that involves bacteria, fungi, algae, or still fraught with danger. Bacteria, in parviruses,” he says. David Bradley
MIP, the plasma is the only load, so even small variations in it change the system, he explains. But the MSP has an added load. “The value of the additional load is well adapted,” he says, “so small changes in the plasma do not have an influence.” To test the device, the researchers performed flow injection cold-vapor (FI-CV) analyses of trace levels of mercury in aqueous solutions. Traditional FI-CV with atomic absorption spectroscopy (AAS) was compared with FI-CV with optical emission spectroscopy (OES), using the MSP as the source. Both methods produced essentially the same results. But FI-CV-AAS is limited to single-element analysis. When the MSP is the atomic emission source, multielement analysis can be performed. “That is one of the biggest advantages,” Broekaert says. The researchers began with FI-CV for mercury because the method is well known and the analyte is of interest for
environmental studies. However, the system could be applied to “all elements which are volatile by themselves or which form volatile compounds,” says Broekaert. “There are many possible applications.” At the moment, however, the researchers are most interested in further miniaturizing the device. It is bigger than it needs to be now, Broekaert says, because they used the materials they had on hand. Only the active region—a 20mm-long channel with a 1-mm-square cross section—must be kept. The rest can be trimmed. Then the researchers will work on integrating the MSP with another device in a single wafer. “We have already developed an operating setup with reduced size and power consumption,” Engel says. “And we think a ready-to-use device could be realized without too much effort.” Elizabeth Zubritsky J A N U A R Y 1 , 2 0 0 0 / A N A LY T I C A L C H E M I S T R Y
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