Analytical Currents: Yeast gives rise to new biosensor

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ANALYTICAL CURRENTS Yeast gives rise to new biosensor University of Washington researchers have taken a bit of yeast, raised the temperature, and watched their new in vivo ligand-binding assay grow by leaps and bounds in the biosensor world. Stanley Fields and Chandra Tucker coupled a ligand-binding domain to a temperaturesensitive version of a metabolic enzyme in the yeast Saccharomyces cerevisiae, creating a modular sensor that recognizes the binding of a ligand to its receptor. As an added bonus, the sensor may be able to quantify ligand-binding affinities. The problems with previous in vivo sensors—made from chimeric proteins, in which a ligand-binding domain is coupled to a “reporter”—have included limited applicability, the possibility of many false positives, and the difficulty of synthesizing chimeric proteins that remain active. The new sensor takes advantage of a convenient property of the enzyme dihydrofolate reductase (DHFR). The mouse version of DHFR (mDHFR) can be expressed in two pieces yet still produce an active enzyme. Furthermore, the researchers found that they could insert a ligand-binding domain between the two pieces of mDHFR and create a

mDHFR

107 108 functional chimeric enzyme. N-DHFR FKBP12 C-DHFR dhFK In some cases, the chimeric enzyme was temperature-sensitive (i.e., X FKBP12 C-DHFR somewhat unstable at 38.5 P66LdhFK P66L °C). This characteristic was quite useful because dhER N-DHFR ERa-LBD C-DHFR the chimeric enzyme could be stabilized by the bindSchematic showing: (top) the mouse version of dihydrofolate ing of a ligand to the ligreductase (mDHFR); a construct in which the two mDHFR doand-binding domain. mains are separated by the ligand-binding domain FKBP12 Thus, the chimeric enand a short linker; the temperature-sensitive version; and zyme’s level of activity in(bottom) a similar construct, which uses the estrogen recepdicated whether or not a tor ER␣ ligand-binding domain. (Adapted with permission. ligand was bound. BeCopyright 2001 Nature America.) cause DHFR is involved in DNA synthesis, the enzyme’s activity level could be determined by measuring the growth human protein FKBP12 and the human estrogen receptor ER␣. In both cases, rate of the yeast population. Therefore, the sensor successfully identified known yeast that bore the chimeric enzyme ligands. Further testing with the estrocould screen a library of compounds gen receptor showed that the yeast popand distinguish the ligands that recogulation growth corresponded to the lignized the ligand-binding domain (good and’s binding affinity, suggesting that population growth) from those that the sensor may be able to quantify ligdidn’t (poor population growth). and binding. (Nat. Biotechnol. 2001, The researchers tested the sensor with two ligand-binding domains: the 19, 1042–1046)

Intelligent automated spectroscopy As part of an effort to develop automated instruments that can hold their own against instruments operated by people, Eric Salin and colleagues at McGill University (Canada) evaluated three classification algorithms for possible use in atomic spectroscopy. Specifically, the researchers tested the algorithms’ ability to differentiate various aluminum and steel alloys. This task was more challenging than some of the problems previously run through

the algorithms. First, the algorithms were tested for their ability to assign a sample to the proper class on the basis of semiquantitative information about the elemental composition—an important task when choosing a calibration methodology. The semiquantitative data were simulated using true values with large (~25%) relative standard deviations (RSDs). Second, the algorithms were tested for their ability to classify samples accurately following quantitative

analysis. In this case, the data were simulated as true values with ~5% RSDs. The Bayesian classification method performed best, achieving 89.8% accuracy with the semiquantitative data and 100% accuracy with the quantitative data. This finding stands in contrast to an earlier study in which the rival C4.5 method had performed well when conditioned with a large training set. (J. Anal. At. Spectrom. 2001,16, 1135–1141)

J A N U A R Y 1 , 2 0 0 2 / A N A LY T I C A L C H E M I S T R Y

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ANALYTICAL CURRENTS Lateral diffusion by SECM What happens when there is no scanning in scanning electrochemical microscopy (SECM)? If you’re Patrick Unwin and colleagues at the University of Warwick and Coventry University (both in the United Kingdom), keeping SECM fixed at a spot means that you can study lateral diffusion in Langmuir monolayers at an air/water interface. To measure lateral diffusion electrochemically, an ultramicroelectrode probe was placed in the water phase just 1–2 µm from the monolayer. A three-step process was then initiated to determine the diffusion kinetics. First, an electroactive species was generated by diffusioncontrolled electrolysis. In this case, 3+ Ru(bipy) 2+ 3 was oxidized to Ru(bipy) 3 . The newly formed electroactive agent diffused to the monolayer where it reacted with an amphiphile at the interface—in this study, N-octadecylferrocenecarboxamide (C18Fc). The now-reacted electroactive species diffused back to the ultramicroelectrode, and the resulting feedback was followed as a current–time transient, which depended on the electron transfer rate between the amphiphile and the

Potential Electrochemical “bleaching” step Red ➞ Ox + ne

Recovery step Ox + ne ➞Red

Analysis step Red ➞ Ox + ne Time

Air Water

Red

Ox

Red

Ox

Red

Ox

One, two, three. The potential step profile and corresponding processes for the three-step scanning electrochemical microscopy measurements of monolayer lateral diffusion at an air/water interface.

agent. Step two was recovery—converting the remaining electroactive agent back to its original form, which in this study meant reducing all the unreacted Ru(bipy) 3+ 3 . Step three—the analysis— simply repeated the first step. Because the amphiphile laterally diffuses during the recovery step, its duration determines the information gathered in the analysis step. If the surface-confined amphiphile is significantly oxidized and the recovery period is short, then

the data for the analysis step will show an inert surface. The tip–interface separation can then be measured. On the other hand, if the recovery period is longer, then lateral diffusion reorganizes the surface—in this example, bringing in unreacted C18Fc. The current–time response is then a function of the amphiphile’s lateral diffusion coefficient, the electron transfer kinetics, and the surface coverage. (J. Phys. Chem. B 2001, 105, 11,120–11,130)

Blue “star” emissions signal benzene Blue luminescent compounds are a natural choice for sensing aromatic compounds because both commonly absorb in the UV or near-UV range. R. Stephen Brown, Suning Wang, and colleagues at Queen’s University (Canada) take this idea a step further and make an unusual

solid-state fluorescent fiber-optic sensor for benzene. The researchers used a tiered approach that began with the creation of several star-shaped organic molecules, which can coordinate with various metal ions. In some cases, the result was a fluorescent sensor complex, also star-shaped, that interacted selectively with other molecules. In particular, a star-shaped complex, formed from the reaction of 1,3,5-tris(p(2,2´-dipyridylamino)phenyl)benzene with ZnCl2 in CH2Cl2/CH3OH, Two views of a star-shaped ZnII-based sensor that detects benzene (yellow). (Adapted with permission. Copyright 2001 Wiley-VCH Verlag GmbH.)

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A N A LY T I C A L C H E M I S T R Y / J A N U A R Y 1 , 2 0 0 2

showed an unusually high selectivity for benzene vapor. To make a fiber-optic sensor, the researchers attached a prepolymerized polydimethylsiloxane bead, which contained the complex, to a fused-silica optical fiber. Exposing the optical fiber to 500-ppm samples of benzene and 12 other compounds, including other aromatic molecules, caused a remarkable decrease in fluorescence intensity with benzene vapors, but not with the other compounds. In addition, X-ray diffraction studies provided some much-needed information on the direct correlation between the sensor’s structure and its capabilities. (Angew. Chem., Int. Ed. 2001, 40, 4042–4045)

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Electronic detection of DNA mismatches Never mind the fancy light show of fluorescent probes. C. J. Yu and colleagues at Motorola Clinical Micro Sensors and the California Institute of Technology have developed a new electrochemical probe for detecting single-base DNA mismatches in an electronic assay. The basic assay works by incorporating a ferrocene-containing phosphoramidite, which provides a range of detectable redox potentials, into an oligonucleotide

probe. A self-assembled monolayer transfers electrons between the ferrocenes and gold electrodes arrayed on a chip. Because mismatch detection requires two probes, one for the wild-type sequence and one for the altered sequence, the researchers fabricated two kinds of probes, each using a different ferrocenyl complex. Using ac voltammetry, the researchers determined that two probes could be distinguished by the positions

of the peaks in the spectra. Furthermore, when the probes were incorporated into a proprietary DNA screening chip, they could identify double-stranded DNA segments in which both strands were wild-type, both were mutated, and the two were mixed—the classic mismatch. The researchers suggest that this assay also could be used to genotype human blood samples. (J. Am. Chem. Soc. 2001, 123, 11,155–11,161)

AC currents (nA)

30

Microfabricated SNOM tips

20

10

0.0 –50

50 150 250 350 450 Potential (mV vs Ag/AgCl)

550

AC currents (nA)

28 24 20 16 12 8.0 4.0 0.0 –50

50

150 250 350 450 Potential (mV vs Ag/AgCl)

550

To avoid some typical problems in fabri-

plete the probe, each guide-tip structure

cating scanning near-field optical mi-

is glued to an optical fiber that has been

croscopy (SNOM) probes, researchers

chemically etched at one end to a diam

such as G. Genolet and colleagues at

of < 50 µm.

IBM’s Zurich Research Laboratory, the

The tips can be batch-fabricated and

University of Neuchâtel, and the Swiss

do not require any subsequent process-

Federal Institute of Technology (all in

ing, such as focused ion-beam milling.

Switzerland) have turned to microfabrica-

The researchers made apertures as small

tion. In particular, this approach is expect-

as 50 ⫻ 130 nm, although problems with

ed to avoid some of the problems that

the tip-to-fiber coupling have so far re-

occur when the tip’s aperture is formed

sulted in poor images for apertures < 300

by the angle evaporation of aluminum.

nm. (Rev. Sci. Instrum. 2001,72, 3877–3879)

The researchers describe the use of standard microfabrication techniques to Fiber core

Optical fiber

make 8.5-µm-tall pyramid-shaped tips with 12-µm bases from UV-curable plas-

AC currents (nA)

20

Etched optical fiber

tic. Pyramid-shaped holes are first etched in a wafer. The apertures at the apexes of

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the tips are defined by etching cavities in

Ring guiding structure

the buried oxide layer beneath the holes.

Metallic coating Tip

A layer of aluminum, which coats the

5.0

Photoplastic part Clearing angle

sides of the pyramids but does not cover 0.0 –50

50 150 250 350 450 Potential (mV vs Ag/AgCl)

550

Alternating current voltammetry spectra for (a) a wild-type DNA sequence, (b) a mutated DNA sequence, and (c) a mismatch between the two.

the apertures, is then deposited. Next, ring-shaped guide structures made from a second, thicker layer (160 µm) of the same plastic are centered over the tips. To com-

Aperture

Schematic of the new scanning near-field optical microscopy probe. (Adapted with permission. Copyright 2001 American Institute of Physics.)


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ANALYTICAL CURRENTS Power up the biosensors Holey dendrimers To organic chemists, dendrimers—the ever-expanding networks of repeating organic moieties—are structures of beauty that offer exciting synthetic challenges. Now, analytical chemists are discovering their practical side. Martin Schlupp, Joachim Bargon, Klaus Müllen, and colleagues at the Universität Bonn and the Max-Planck-Institut für Polymerforschung (both in Germany) show that a quartz microbalance (QMB) surface coated with one type of dendrimer can be an effective detector for volatile organic compounds (VOCs). The researchers used polyphenylene dendrimers, which have rigid frameworks and fixed internal voids, and studied one series with different

Although most fuel cell research is aimed at high energy and efficiency, Itamar Willner and his co-workers at the Hebrew University of Jerusalem (Israel) took exactly the opposite tack. Low efficiency for their biofuel cells translates into highly specific information about the concentration of an analyte. These self-powered biosensors put out enough energy (1 µW) for a good signal but not enough to cause contaminating reactions that would clutter their data. The sensor is essentially a flow cell made from two electrodes separated by a spacer. The anode is functionalized with an enzyme layer—in one version of the biosensor, glucose oxidase. To form an effective electrical contact between the glucose oxidase and the electrode, the researchers use a previously reported process in which the apoenzyme is reconstituted on a flavine adenine dinucleotide layer, which is covalently linked to the electrode. The enzyme catalyzes

the oxidation of glucose—the analyte or fuel—to gluconic acid. This yields lowlevel electrical power, and the open-circuit voltage of the system is determined by the fuel concentration. The cycle is complete with the simultaneous reduction of oxygen to water on a cathode functionalized with cross-linked cytochrome c/cytochrome oxidase. A second version of the sensor replaces glucose oxidase with lactate dehydrogenase using a similar functionalization process. The sensors could detect glucose and lactate, respectively, in the 1- to 80-mM range. The cells were operated continuously for 5–7 h. Because the cells are self-powered by biological fluids, Willner and colleagues suggest that they may be used as invasive sensing devices. The researchers also say that the sensors can be tailored to detect substances such as alcohols, fructose, and amino acids. (J. Am. Chem. Soc. 2001, 123, 10,752–10,753)

core organic moieties and another that inlet

was based on a core moiety with variAu

ous functional groups. The dendrimers

Spacer Glass

were electrosprayed onto the QMBs, and the responses to various VOCs

A e-

were measured. As expected, doubling the surface coating increased

S

H N PQQ

FAD Gluconic acid

GOx

the signal 2-fold. Moreover, the dendrimers were stable for months, re-

Glucose

e-

O2

COx

sponded quickly, and were reusable. The dendrimer series built from dif-

H2O

B

fering core moieties all showed selec-

S

tivity to polar VOCs, such as benzaldehyde, nitrobenzene, and acetophenone.

S

The functionalized dendrimers—constructed with 16 cyano, carboxyl, or

Au

C

e-

Au

S

H N PQQ

NAD+ COx

eCyt c Cys S O

LDH H N PQQ eH N PQQ

NAD+ e-

Cyt c Cys S

Lactate

eLDH

NAD+

O N O O

N

O N H

S

O N H

S

Pyruvate

imino groups—had more varied responses, with the carboxyl dendrimer detecting acetone, isopropyl methyl ketone, and nitromethane. (Angew. Chem., Int. Ed. 2001,40, 4011–4015)

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outlet

Self-powered biofuel cell biosensors with anodes that detect either (A) glucose or (B) lactate. Both sensors use (C) the cytochrome c/cytochrome oxidase-functionalized cathode.


Au

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RESEARCH PROFILES Herding chromosomes down the chiseled trail other at 120°. The top surface is a cover long chain, so it cannot pass back over Imagine a setting where double-strandthe fence. Instead, it concentrates in a slip treated to improve wetting with the ed DNA in its uncleaved form is free to narrow band next to the barrier. Finally, mobile phase. This microscopic topoloreptate through an open gel. Stripped the voltage is reversed again to push the gy forms the foundation of a gel with of the protein coat that protects DNA narrow band downstream into the oblarge canyons of space at very prein vivo, the molecule is corralled at a stacle course. dictable orientations and a porosity imvery narrow starting point and driven Within the obstacle course, more hopossible to achieve in classic agarose back and forth through chutes wide mogeneous electric fields enough to allow a chromodrive the DNA from side to some to unravel. In this kind side. “This see-saw motion is of “DNA drive” (see figure), essential to getting the DNA unfurled DNA travels at a futo unfurl,” says Austin. And rious pace—a constant pace having the DNA unfurled is inversely proportional to its the key to performing the extended length—in a landseparation based on length. scape carved out of quartz (To see a movie of the polyand immersed in an ever-remer dynamics in a sculpted versing electric field. gel, visit http://suiling. In the December 15 issue princeton.edu/hexagonal/ of Analytical Chemistry (pp hex.html.) 6053–6056), Robert Austin, Periodically switching the Olgica Bakajin, and a team of direction of the electric field others from Princeton Uniacross the direction of travel versity and Cavendish Labois routine in standard agarose ratory (United Kingdom) degels. However, classic gels scribe such a device, which is are so congested that only essentially an obstacle course the smaller DNA fragments tailored to streamline DNA can travel into the gel, and DNA dressed up as “blue genes” proceed in a sidewinder stampede separation. The researchers they do that very slowly. across miniature “badlands” in this false color micrograph. resolve DNA molecules of Standard gels can take ~50 and ~170 kilobase pairs hours—even days—to run, (kbp) from each other in just and longer strands of DNA are so hin10 s. Just 20 min is enough time to sep- gels. By switching the direction of the dered in the gel that they are slowed to electric field back and forth across the arate up to 20 bands of DNA spanning impractical speeds. The open nature of separation axis, DNA strands of ~165 these lengths on a hexagonal device ~3 the sculpted quartz “gels” eliminates kbp can be teased out into long threads cm in diam. this problem and, as a bonus, makes it and transit 1 cm of the array in 11 min. “We chose these example DNA molpossible to calculate the lengths of mol“One key to achieving rapid resoluecules because they were large and comecules on the basis of the applied voltmercially available.” says Austin. But the tion of DNA by size is to start with the age waveform and the rate of travel. device makes it possible to separate much whole collection in one narrow band,” Another advantage is that the devices larger pieces of DNA, and probably whole says Austin. To achieve this, Austin’s group uses entropic focusing, which was don’t have to be made from quartz. chromosomes, in assays that are much developed by Harold Craighead’s group “We can use the quartz devices as masfaster than the classic electrophoresis ters and generate plastic ones from at Cornell University. gels, he adds. them,” says Austin. Plastic devices The researchers begin by applying a The sculpted, two-dimensional hexwould be less expensive and easier high potential, which enables the DNA agonal array of silicon dioxide pillars is to mass-produce. formed on a quartz substrate by carving to deform and pass over a long, narrow Ultimately, Austin’s goal is to increase out the space in between using standard “fence”. Then the researchers reverse the polarity and decrease the voltage to the speed of routine genetic screening microlithography techniques. The pildrive the DNA back toward the barrier. of entire organisms. I suggest you hold lars, 2-µm tall, 2-µm wide, and 2-µm At low voltages, DNA lacks the internal on to your hat. a apart, describe three sets of thousands –Zelda Ziegler energy required to order itself into a of parallel, open channels crossing each


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MEETING NEWS 5th International Conference on Miniaturized Chemical and Biochemical Analysis Systems (µTAS 2001)—Elizabeth Zubritsky reports from Monterey, CA

News from the

Problem-solving microplasma If you think about plasmas, you might picture plasma-based MS or optical emission detection. You probably don’t think, “Great for solving computational problems!” But Andreas Manz and Darwin Reyes at Imperial College (United Kingdom) and George Whitesides at Harvard University have found that plasmas are very useful for finding the shortest route between two points—a type of combinatorial optimization problem. The researchers started with a simple maze etched into glass chips. Tungsten wires marked the starting and ending points. Helium gas was fed through a capillary into the covered chip, and a voltage was applied. The resulting glow discharge followed the shorter of two possible paths through the maze. A more complex chip recreated the layout of central London streets in a network of etched channels, 250-µm wide and 100-µm deep. A wire marked the starting point, Imperial College, and an ending point of either Trafalgar Square, Buckingham Palace, or Victoria Station. Again, the glow discharge mapped the shortest route between the points. “One of the nicest features is that the answer . . . is shown in a visible way, without any need for translation,” Reyes says. “You just see the path glowing.” A variation on this theme is to operate above the breakdown voltage. So far, applying a voltage equal to the breakdown voltage has always resulted in the shortest path, Reyes says. But if the voltage is increased, the glow discharge finds alternate paths—a phenomenon that might be useful for solving problems such as rerouting traffic, he explains. From the viewpoint of the computing community, however, the microplasma approach is limited, Reyes says. The system doesn’t perform mathematical operations, for example, and it isn’t a generalpurpose computational tool. Nevertheless, an advantage of the sys-

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tem is that the time needed to find the shortest path is relatively independent of the complexity of the network, Reyes says. The complexity increases quickly with the number of nodes, or decision points. If each node has two options, then a network with 11 nodes has 211 possible paths; the central London map had 2456 possible paths. Although the researchers have not yet conducted a thorough investigation of the relationship, they have learned that increasing the number of nodes 90-fold merely doubles the amount of time needed to find a solution.

wants to make a series of stops. To do this, the researchers won’t be able to rely on wires to define the stops, as they do now, Reyes says. Instead, they will need to find a way to manipulate the plasma, perhaps with electric or magnetic fields. Another test is finding the shortest path around a series of obstacles in threedimensional space. This is the type of navigational problem that robot designers face, and it is more difficult than map navigation because the system has fewer constraints, Reyes says. The preliminary data here are promising, but he is not ready to say that the system has solved this problem. As exciting as all of the results to date are, Reyes is cautious. “The early work was easy,” he says, “because you knew where you wanted to start and where you wanted to end.” The system’s real potential will be determined by its performance on the truly difficult problems.

Better turns for microchannels A glow discharge highlights the shortest route through a maze. (Adapted with permission from Ramsey, J. M.; van den Berg, A. Micro Total Analysis Systems; Kluwer Academic Publishers: Dordrecht, The Netherlands, 2001; pp 37–39.)

The performance might be further enhanced by miniaturizing the system even more, Reyes adds. Because the pathlengths would be shorter, such a reduction should lower the voltage required (15–21 kV) and reduce the time needed to find a path through the chip. It would also squeeze more nodes—perhaps as many as 1000—into the same area. The researchers have already begun to tackle more challenging problems. One is solving the classic combinatorial optimization problem of the traveling salesman. In this case, the goal is to plot the shortest route for a salesman who


Adding a bend to a microchannel may mean taking a turn for the worse, because turns often distort nice, neat sample bands. Researchers have devised various solutions to this problem of “band dispersion” (Anal. Chem. 2000, 72, 687 A–690 A), and now, Stewart Griffiths and Robert Nilson at Sandia National Laboratories–Livermore describe a general numerical method for optimizing turns and junctions, such as Ts and Ys (Anal. Chem. 2001, 73, 272–278). The resulting geometries work for electroosmosis, electrophoresis, and some pressure-driven flows, the researchers say. Turns are troublesome because the part of the band that travels along the inside wall of the channel generally winds up ahead of the part that travels along the outside wall—a phenomenon sometimes called the “racetrack effect”. In electrically driven flows, this problem may be exacerbated by variations in the electric field. People sometimes assume that equal-

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izing the pathlengths along the inner and outer channel walls will fix the problem, notes Griffiths. But it’s really “an observation after the fact” that a good solution equalizes the pathlength, he says. It isn’t the main source of the improvement. For example, the inner channel wall’s length could be increased by adding a small sine wave, but this would do little to correct the band dispersion. The strategy used by Griffiths and Nilson is to counter-rotate the sample band by narrowing and expanding the turn asymmetrically—bringing the inner wall closer to the outer wall. The counter-rotation induced by the asymmetry at the beginning and end of the turn largely corrects the rotation that usually occurs in the rest of the turn. Griffiths notes that the asymmetry in the channel constriction is the source of the counter-rotation. Narrowing the channel symmetrically or moving the outer wall instead would not have the same effect. The optimized Ts and Ys work on the same principle of contracting and expanding the channel to produce a counter-rotation. However, this approach induces an unwanted side effect: The sample band becomes bowed by channel expansions and contractions, notes Griffiths. Surprisingly, expansions and contractions don’t cancel one another; the band is bowed in the same direction in both cases. Therefore, their designs make a tradeoff between correcting the racetrack effect and not inducing too much bowing.

(a)

(b)

(a) An ordinary hairpin turn exhibits considerable band dispersion. (b) An optimized turn, which uses an asymmetrical taper, induces little distortion.

Modeling electrokinetic transport is surprisingly difficult, Griffiths observes. On the one hand, the researchers have the unusual luxury of working with a well-defined system of equations that govern flow and species transport in microchannels, he says. “[But] you cannot solve those equations for any real system of interest,” he adds. “It’s just too computationally demanding.” That’s why people solve the equations for one component—for example, a turn or a T—at a time. But even then, researchers make simplifications and approximations. In addition, the turn problem can’t be solved in the manner of a typical optimization, Griffiths explains. Normally, the approach would be to represent the inner and outer channel walls mathematically as two series of boundary points. An optimization algorithm would move the points one at a time and, in each case, determine whether the variance of a

band positioned downstream would increase or decrease. Eventually, this process would produce a solution better than the initial guess. But for this problem, says Griffiths, “Every change you make in the boundary—if you make the changes one point at a time—makes the variance go up.” In a sense, every change makes the boundary “rougher”, and the algorithm wanders around, unable to find a better solution. “We never really got to the bottom of why [that] would be,” he adds. But they got around the issue and obtained successful optimizations by representing the boundary using a set of critical parameters instead of points. Because the researchers’ numerical method is so general, they have received requests to make the software available to everyone. But Griffiths says that the code isn’t in a neat, user-friendly package that he can distribute.

GOVERNMENT AND SOCIETY Analysis comes to dietary supplements For years, U.S. law has allowed dietary supplements to be sold without most of the oversight required for conventional medicines, including a rigorous quality assurance/quality control (QA/QC) accounting of ingredients. Starting last fall, AOAC International (formerly the Association of Official Analytical Chemists),

the U.S. Food Drug Administration (FDA), and the National Institutes of Health’s Office of Dietary Supplements (NIH-ODS) began developing QA/QC laboratory methods for the dietary supplements containing ephedrine-type alkaloids and aristolochic acid. These are the first dietary supplements

for which standardized laboratory methods have been developed, according to AOAC. The supplements are of particular interest because they can cause health problems. Herbal and synthetic ephedrinetype alkaloids—which include the active compounds norephedrine, ephedrine, pseudoephedrine, and methylephedrine—


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GOVERNMENT AND SOCIETY are found in popular items such as energy-boosting sports drinks, weight-loss formulas, and upper respiratory medications. The side effects vary and include increased blood pressure, insomnia, anxiety, and muscle disturbances. Aristolochic acid, a contaminant in some Chinese herbal medicines, can cause renal failure. The new methods are just the first step in regulating and ensuring the safety of dietary supplements, says James Bradford of AOAC. However, neither AOAC nor NIH-ODS will say which supplements might come next. Although the federal budget has not been finalized, it appears that Congress will appropriate money to NIH-ODS to develop these analytical methods, say the experts. This allocation of funds is largely the result of lobbying from the dietary supplement community, according to Leila Saldanha at the Consumer

Healthcare Products Association. The industry wants methods that will hold up in a court of law as tools for enforcement and product competitiveness. The industry also wants to establish Good Manufacturing Practice standards to gain consumer confidence in the safety of the products, says Paul Coates at NIH-ODS. Coates adds that standardized and validated laboratory methods will also be important for NIH-ODS. The office is generally interested in the safety and quality of dietary supplement products and plans to run clinical trials to assess the safety of ephedrine-type alkaloids. Since 2000, the AOAC Dietary Supplement Task Group—a collaboration between federal agencies such as FDA, five trade organizations, the National Research Council of Canada, U.S. Pharmacopeia, and the National Institute of Standards and Technology—has been reviewing

the methodologies for the ephedrinetype alkaloids and aristolochic acid and researching other dietary supplements. Two methods based on LC—one using MS as the detector and the other relying on fluorescence or UV measurements—will be developed to determine the concentration of ephedrine alkaloids in supplements. The laboratory methods will cover various matrixes, including botanical raw materials, extracts, complex mixtures of multiple dietary supplements, high-protein drinks, and animal specimens. Different sample matrixes may require more than one testing method, says Anita Mishra-Szymanski of the AOAC Official Methods/Peer Verified Methods Program. For aristolochic acid, AOAC plans to validate an LC/MS method currently used by FDA. Method development and validation is expected to take 18 months. a –Laura Ruth

on a leave of absence). His research interests and expertise include the development and application of methods for chemical and physical measurements, industrial analysis, and quality assurance, including certification of reference materials and interlaboratory comparisons.

from James Madison University, her M.S. in bioengineering from the University of Utah, and her Ph.D. in toxicology from the University of Maryland Medical School–Baltimore. Her research interests include the application of liposomes in analytical measurements and the development and application of microfluidics, biosensors, and integrated sensor systems.

PEOPLE New Advisory Board and A-Page Advisory Panel members appointed Six new members from government, academia, and industry have been selected to serve three-year terms on Analytical Chemistry’s Editorial Advisory Board. Established in the 1940s, the board is a vital link between the Journal editors and the analytical chemistry community, providing guidance and advice on editorial content and policy.

Advisory Board Manfred Grasserbauer, director of the Institute for Reference Materials and Measurements (Belgium), received his degree in chemistry and a doctorate of technical sciences from the Technische Universität Wien (Austria), where he is also a professor (currently

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Michele Kelly, project leader at Pfizer Global R&D, received her B.S. from James Madison University and her Ph.D. from the University of Maryland–Baltimore County. Her research interests include the application of MS for the analysis of chemical leads and biological targets in drug discovery. Laurie E. Locascio, project leader in the analytical chemistry division of the National Institute of Standards and Technology, received her B.S. degree in chemistry


Richard L. McCreery, professor of chemistry at Ohio State University, received his B.S. in chemistry from the University of California–Riverside and his Ph.D. from the University of Kansas. In his research, he uses spectroscopic probes of electrochemical processes and molecular elec-

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tronics in an effort to relate surface structure to behavior. Susan Olesik, professor at Ohio State University, received her B.A. from DePauw University and her Ph.D. from the University of Wisconsin–Madison. Her research interests include fundamental studies of separation science, supercritical fluid and high-fluidity liquid technology, and the development of carbon-based surfaces for sensors and separation science applications. Steve Weber, professor at the University of Pittsburgh, received his B.A. from Case Western Reserve University and his Ph.D. from McGill University (Canada). His research interests include sensors and selective extraction using molecular recognition, peptide determinations, electrochemistry, band spreading in separations, and post-column reactions.

A-Page Advisory Panel Analytical Chemistry has also selected five new members to serve three-year terms on its A-Page Advisory Panel. The panel provides feedback on the A-page editorial content and proposes appropriate topics and authors for feature articles. Yves Guillaume, professor of analytical and bioanalytical chemistry at the University of Franche-Comté (France), received his Ph.D. in both pharmacy and analytical–physical chemistry. His research interests include chemometrics, the mechanisms of separation in chromatography and electrophoresis, guest–host interactions, nonequilibrium

chromatographic techniques, and applying separation analysis methods to environmental and biological problems, including new transport systems for pharmacomolecules. Steve A. Hofstadler, executive director of drug discovery in the Ibis Therapeutics division of Isis Pharmaceuticals, Inc., received his Ph.D. from the University of Texas–Austin. His research interests include the development and application of MS-based bioanalytical techniques, including highthroughput FT-ICR methodologies to characterize noncovalent complexes of nucleic acids, proteins, and small molecules. Sharon L. Neal, associate professor at the University of Delaware, received her Ph.D. from Emory University. Her research interests include the development of multidimensional fluorescence measurements that integrate traditional photophysical and photodynamic measurements. Her recent work includes the application of

such methods to biomembrane characterization and studies of small molecule–biomolecule interactions. Carol L. Nilsson, associate professor at Göteborg University’s Institute of Medical Biochemistry (Sweden), received her M.D. and Ph.D. from that university. Her research interests include biomedical applications of MS and proteomics, especially in the study of the expression and structure of membrane proteins, glycoproteins, and protein–carbohydrate interactions. Peter Roepstorff, a professor in protein chemistry at the University of Southern Denmark–Odense, received two candidate degrees: one in physiological chemistry from the University of Aix-Marseille (France) and the other in chemical engineering from the Danish Technical University. His research interests are in protein chemistry, protein MS, and proteomics. In addition, he has made contributions to mass spectrometric analyses of carbohydrates and nucleic

BUSINESS PerkinElmer acquires Packard BioScience In late fall, PerkinElmer moved deeper into the drug discovery market by acquiring Packard BioScience, a provider of drug discovery tools. The merger adds automated liquid-handling, samplepreparation, and biochip technologies to PerkinElmer’s presence in the instrumentation, life sciences, and optoelectronics fields. Kevin Lorenc, director of corporate communications for PerkinElmer, says that life sciences, particularly in the drug

discovery market, is the fastest growing area of the company. “Last year [2001],” says Lorenc, “the drug discovery market made up about 75% of the revenue for the life sciences division, with genetic disease screening making up the other 25%.” To capitalize on both of these areas, the company has created the PerkinElmer Life Sciences’ Drug Discovery division, headed by Frank Witney, former president of Packard BioScience. The new division expects revenue in 2002 to be in the range of $525 million dollars. a –Wilder D. Smith


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