Diffusion coefficient measurement in a microfluidic analyzer using dual-beam microscale-refractive index gradient detection. Application to on-chip molecular size determination

We report a microchip-based detection scheme to determine the diffusion coefficient and molecular mass (to the extent correlated to molecular size) of analytes of interest. The device works by simultaneously measuring the refractive index gradient (RIG) between adjacent laminar flows at two different positions along a microchannel. The device, referred to as a microscale molecular mass sensor (micro-MMS), takes advantage of laminar flow conditions where the mixing of two streams occurs essentially by diffusion across the boundary between the two streams. Two flows merge on the microchip, one containing solvent only, referred to as the mobile phase stream and one which contains the analyte(s) of interest in the solvent, i.e. the sample stream. As these two streams merge and flow parallel to each other down the microchannel a RIG is created by the concentration gradient. The RIG is further influenced by analyte diffusion from the sample stream into the mobile phase stream. Measuring the RIG at a position close to the merging point (upstream signal) and simultaneously a selected distance further down the microchannel (downstream signal) provides real-time data related to the extent a given analyte has diffused, which can be readily correlated to analyte molecular mass by taking the ratio of the downstream-to-upstream signals. For the dual-beam RIG measurements, a diode laser output is coupled to a single mode fiber optic splitter with two output fibers. Light from each fiber passes through a graded refractive index (GRIN) lens forming a collimated beam that then passes through the microchannel and then on to a position sensitive detector (PSD). The RIG at both detection positions deflects the two collimated probe beams. The deflection angle of each beam is then measured on two separate PSDs. The micro-MMS was evaluated using polyethylene glycols (PEGs), sugars, and as a detector for size-exclusion chromatography (SEC). Peak purity can be readily identified using the micro-MMS with SEC. The limit of detection was 0.9 ppm (PEG at 11 840 g/mol) at the upstream detection position corresponding to a RI limit of detection (LOD) (3sigma) of 7-10(-8) RI. The pathlength for the RIG measurement was 200 microm and the angular LOD was 0.23 micro(rad) with a detection volume of 8 nl at both positions. The average molecular mass resolution was 9% (relative standard deviation) for a series of PEGs ranging in molecular mass from 106 to 22 800 g/mol. With this excellent mass resolution, small molecules such as monosaccharides, disaccharides, and so on, are readily distinguished. The sensor is demonstrated to readily determine unknown diffusion coefficients.     

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Characterization and complementation of a Pichia stipitis mutant unable to grow on d-xylose or l-arabinose

Pichia stipitis CBS 6054 will grow on d-xylose, d-arabinose, and l-arabinose. d-Xylose and l-arabinose are abundant in seed hulls of maize, and their utilization is important in processing grain residues. To elucidate the degradation pathway for l-arabinose, we obtained a mutant, FPL-MY30, that was unable to grow on d-xylose and l-arabinose but that could grow on d-arabinitol. Activity assays of oxidoreductase and pentulokinase enzymes involved in d-xylose, d-arabinose, and l-arabinose pathways indicated that FPL-MY30 is deficient in d-xylitol dehydrogenase (D-XDH), d- and l-arabinitol dehydrogenases, and d-ribitol dehydrogenase. Transforming FPL-MY30 with a gene for xylitol dehydrogenase (PsXYL2), which was cloned from CBS 6054 (Gen Bank AF127801), restored the D-XDH activity and the capacity for FPL-MY30 to grow on l-arabinose. This suggested that FPL-MY30 is critically deficient in XYL2 and that the d-xylose and l-arabinose metabolic pathways have xylitolas a common intermediate. The capacity for FPL-MY30 to grow on d-arabinitol could proceed through d-ribulose.     

Anionenaustauschtrennungen der mit Tributylphosphat extrahierbaren Elemente. VII

Zusammenfassung Eine Methode zur Abtrennung des Urans wird beschrieben, bei der das Uran aus einer 6-m salzsauren Lösung mit reinem, unverdünntem TBP extrahiert und aus einer Mischung von 30 Vol.% TBP, 60 Vol.% Methylglykol und 10 Vol.% 12-m Salzsäure am stark basischen Anionenaustauscher Dowex 1-X8 (Chloridform) adsorbiert wird. Da der TBP-Extrakt zur Bereitung dieser Mischung verwendet wird, lassen sich Extraktion und Ionenaustausch kombinieren, wodurch nicht nur die Selektivität der Abtrennung des Urans wesentlich erhöht wird, sondern auch eine so weitgehende Anreicherung des Urans auf dem Harz erfolgt, daß auch ppm-Mengen leicht zu bestimmen sind. Das Harz wird zu diesem Zweck zunächst zur Entfernung des TBP mit einer Mischung von 90 Vol.% Methylglykol und 10 Vol.% 12-m Salzsäure, dann zur Entfernung von mitadsorbierten Elementen mit 6-m Salzsäure behandelt; anschließend wird das Uran mit 1-m Salzsäure eluiert und fluorimetrisch bestimmt. Zu Vergleichszwecken wurden 16 geologische Proben nach dieser Methode und auch unter Anwendung einer anderen Anionenaustauschmethode analysiert. Die Ergebnisse zeigen, daß das hier beschriebene Trennverfahren eine quantitative Abtrennung des Urans ermöglicht und zu gut reproduzierbaren Resultaten führt.

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Anionic exchange separations of the elements that are extractable with tributyl phosphate. VII

Summary A method is described for the separation of uranium in which the latter is extracted from a 6M hydrochloric acid solution by means of pure undiluted TBP and adsorbed from a mixture of 30 volume % TBP, 60 volume % methylglycol and 10 volume % of 12M hydrochloric acid on the strongly basic anionexchanger Dowex 1-X8 (chloride form). Since the TBP-extract is used for the preparation of this mixture, the extraction and the ion-exchange may be combined, whereby not only the selectivity of the separation of the uranium is significantly raised but also there follows such a rar-reaching enrichment of the uranium on the resin that even ppm quantities are readily determined. For this latter purpose, the resin is first treated with a mixture of 90 volume % methylglycol and 10 volume % 12M hydrochloric acid to remove the TBP, and then with 6M hydrochloric acid to remove the co-adsorbed elements; and then the uranium is eluted by means of 1M hydrochloric acid and determined fluorimetrically. For comparison purposes, 16 geologic specimens were analyzed in accord with this procedure and also employing another anion exchange method. The results show that the separation method described here makes possible a quantitative separation of the uranium and leads to good reproducible results.