Bioselective Membrane Electrode Probes

Author(s): Garry A. Rechnitz

Source: Science , Oct. 16, 1981, New Series, Vol. 214, No. 4518 (Oct. 16, 1981), pp. 287-291
Published by: American Association for the Advancement of Science

Stable URL: https://www.jstor.org/stable/1686863

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14. W. F. Haddon, in High Performance Mass 35. A. F. Weston, K. R. Jennings, S. Evans, R. M. Metcalf, and G. R. Hill, paper presented at the
Spectrometry, M. L. Gross, Ed. (American Elliott, Int. J. Mass Spectrom. Ion Phys. 20, 317 ASMS meeting, Minneapolis, May 1981.
Chemical Society, Washington, D.C., 1978), pp. (1976). 56. W. F. Haddon and R. Molyneux, paper present-
97-1 19. 36. B. Shushan and R. K. Boyd, Anal. Chem. 53, ed at the MS-MS Conference, Asilomar, Calif.,
15. F. W. McLafferty and T. A. Bryce, J. Chem. 421 (1981). September 1980.
Soc. Chem. Commun. 1967, 1215 (1967). 37. G. J. Louter, A. J. H. Boerboom, P. F. M. 57. V. J. Caldecourt, D. Zackett, J. C. Tou, paper
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R. Lester, Metastable Ions (Elsevier, Amster- Mass Spectrom. Ion Phys 33, 335 (1980). May 1981.
dam, 1973). 38. F. W. McLafferty and P. J. Todd, Org. Mass 58. M. Linscheid, J. D'Angona, A. L. Burlingame,
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kataraghavan, F. W. McLafferty, J. Am. Chem. 39. F. W. McLafferty, P. J. Todd, D. C. McGilvery, U.S.A. 78, 1471 (1981).
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18. D. H. Smith, C. Djerassi, K. H. Maurer, U. (1980). I. J. Amster, C. J. Sack, F. W. McLafferty,
Rapp, ibid. 96, 3482 (1974). 40. R. A. Yost, paper presented at the Summer paper presented at the ASMS meeting, Minne-
19. T. L. Kruger, J. F. Litton, R. W. Kondrat, R. G. Symposium, Analytical Division, American apolis, May 1981.
Cooks, Anal. Chem. 48, 2113 (1976). Chemical Society, Pittsburgh, Pa., July 1981. 60. H. R. Morris, M. Panico, M. Judkins, A. Dell,
20. R. W. Kondrat and R. G. Cooks, ibid. 50, 81A 41. W. R. Davidson, J. Fulford, N. M. Reid, T. R. McDowell, paper presented at the ASMS
(1978). Sakuma, B. Shushan, B. A. Thomson, paper meeting, Minneapolis, May 1981.
21. R. A. Yost and C. G. Enke, ibid. 51, 1251A presented at the American Society of Mass 61. J. L. Occolowitz, M. P. Barbalas, F. W. McLaf-
(1979). Spectrometry (ASMS) meeting, Minneapolis, ferty, paper presented at the ASMS meeting,
22. F. W. McLafferty, Accounts Chem. Res. 13, 33 Minn., May 1981. New York, May 1980.
(1980). 42. VG-Micromass, Altrincham, Cheshire, En- 62. E. J. Gallegos, Anal. Chem. 48, 1348 (1976).
23. T. H. Maugh II, Science 209, 675 (1980). gland. 63. D. Zakett, A. E. Schoen, R. W. Kondrat, R. G.
24. P. F. Bente, III, and F. W. McLafferty, Pract. 43. Kratos, Ltd., Urmston, Manchester, England. Cooks, J. Am. Chem. Soc. 101, 6783 (1979).
Spectrosc. 3, 253 (1980). 44. Sciex, Inc., Thornhill, Ontario, Canada. 64. B. Shushan, N. J. Bunce, R. K. Boyd, C. T.
25. J. R. B. Slayback and M. S. Story, Ind. Res. 45. Finnigan Corp., Sunnyvale, Calif. Corke, Biomed. Mass Spectrom. 8, 225 (1981).
Dev. 1981, 129 (February 1981). 46. Extranuclear, Inc., Pittsburgh, Pa. 65. F. W. McLafferty, A. Hirota, M. P. Barbalas,
26. K. Levsen, Adv. Mass Spectrom. 8, 897 (1980). 47. G. L. Glish, D. Zackett, P. H. Hemberger, R. G. R. F. Pegues, Int. J. Mass Spectrom. Ion Phys.
27. D. F. Hunt, G. C. Stafford, Jr., F. W. Crow, J. Cooks, paper presented at the ASMS meeting, 35, 299 (1980).
W. Russell, Anal. Chem. 48, 2098 (1976). New York, May 1980. 66. F. W. McLafferty, A. Hirota, M. P. Barbalas,
28. P. Price, D. P. Martinsen, R. A. Upham, H. S. 48. R. T. McIver, personal communication. Org. Mass Spectrom. 15, 327 (1980).
Swofford, Jr., S. E. Buttrill, Jr., ibid. 47, 190 49. B. S. Freiser, personal communication. 67. G. M. Pesyna, R. Venkataraghavan, H. E.
(1975). 50. W. F. Haddon and F. W. McLafferty, Anal. Dayringer, F. W. McLafferty, Anal. Chem. 48,
29. E. L. Horning, M. G. Horning, P. I. Carroll, I. Chem. 41, 31 (1969). 1362 (1976).
Dzidic, R. N. Stillwell, ibid. 45, 936 (1973). 51. D. Zackett, A. E. Schoen, R. G. Cooks, P. H. 68. P. J. Todd and G. L. Glish at Oak Ridge
30. F. W. McLafferty, P. F. Bente, III, R. Kornfeld, Hemberger, J. Am. Chem. Soc. 103, 1295 National Laboratory have used secondary ion
S.-C. Tsai, I. Howe, J. Am. Chem. Soc. 95, 2120 (1981). bombardment for MS-MS analysis of samples of
(1973). 52. D. F. Hunt, J. Shabanowitz, A. B. Giordani, low volatility (personal communication).
31. F. W. McLafferty, Philos. Trans. R. Soc. Lon- Anal. Chem. 52, 386 (1980). 69. Discussions with I. K. Mun and M. P. Barbalas
don Ser. A 293, 93 (1979). 53. D. F. Hunt, J. Shabanowitz, A. B. Giordani, T. were particularly helpful in preparing this arti-
32. M. S. Kim and F. W. McLafferty, J. Am. Chem. M. Harvey, paper presented at the ASMS meet- cle. The generous financial support of the MS-
Soc. 100, 3279 (1978). ing, Minneapolis, May 1981. MS research program at Cornell by the National
33. P. J. Todd and F. W. McLafferty, Int. J. Mass 54. J. A. Chakel, C. A. Myerholtz, C. G. Enke, Institutes of Health (grant GM16609) and the
Spectrom. Ion Phys. 38, 371 (1981). paper presented at the ASMS meeting, Minne- Army Research Office, Durham (grant
34. A. R. Hubik, P. H. Hemberger, J. A. Laramee, apolis, May 1981. DAAG29-79-C-0046), is gratefully acknowl-
R. G. Cooks, ibid. 102, 3997 (1980). 55. H. L. C. Meuzelaar, W. H. McClennan, G. S. edged.

bilities of membrane electrodes to bio-

logical materials through the use of en-
zyme catalysis to convert substrates to
species that could be sensed by ion-
selective membrane electrodes. The re-
sulting "enzyme electrodes" represent
Bioselective Membrane an increasingly practical, but now largely
conventional, means of effecting biose-
Electrode Probes lective measurements with membrane
electrodes (11).
A special impetus was given to re-
Garry A. Rechnitz search on bioselective membrane elec-
trodes in the early 1970's when stable
and reliable potentiometric sensors for
ammonia, carbon dioxide, hydrogen sul-
One of the most rapidly expanding Potentiometric membrane electrodes fide, and other dissolved gases became
commercially available on a routine ba-
research areas relating to analytical mea- have their conceptual origin in the ubiq-
surements is the development of poten- uitous pH glass membrane electrode sis. Such electrodes combine the tech-
tiometric membrane electrodes with se- which, in its present form, is still the nology of ion-selective membrane elec-
lectivity for ions, dissolved gases, and most sensitive and selective of all such trodes with that of microporous synthet-
biological materials. Activity in this fieldelectrodes. During the past 20 years, a ic membranes (12). The ammonia gas-
is so intense that new publications are wide variety of conventional potentio- sensing membrane electrode, for exam-
appearing at a rate approaching 500 per metric ion-selective membrane elec- ple, is a potentiometric sensor in which a
year (1). Entirely aside from the appeal- trodes based on glasses (4), crystals (5), hydrophobic gas-permeable membrane
ing practical possibilities for such poten- and various liquid membranes (6-8) has is superimposed on a flat pH-type glass
tiometric membrane electrodes, it ap- been developed and commercialized. membrane electrode in contact with a
pears that new research directions have Several recent reviews (1) and mono- thin layer of ammonium chloride electro-
been directly stimulated by the timely graphs (9, 10) give comprehensive ac- lyte solution. This arrangement gives ex-
infusion of concepts from physics (2) counts of this work. ceptional selectivity for the measure-
and biology (3). Some of the conse- Throughout the period of ion-selective
quences of the latter are examined in this electrode development, efforts were
The author is Unidel Professor of Chemistry,
article. made to extend the measurement capa- University of Delaware, Newark 19711.

SCIENCE, VOL. 214, 16 OCTOBER 1981 0036-8075/81/1016-0287$01.00/0 Copyright ? 1981 AAAS 287

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ment of ammonia in the presence of ions, eration at 4?C for months without loss of useful lifetimes owing to deterioration of
biological molecules, and even other dis- activity. the tissue layer. This does not occur if
solved gases. It will be seen below that In phosphate buffer at pH 7.8, the- a simple preservative such as sodium
these gas-sensing electrodes are key ele- resulting electrode gives a response of azide is added to the working buffer. In
ments in the development of the newer approximately 50 millivolts per tenfold the absence of the preservative, the tis-
types of bioselective probes. glutamine concentration change over the sue electrode shows a loss of response
range of 6.0 x 10-5M to 6.7 x 10-3M and selectivity; eventually, bacterial
glutamine, with a lower limit of detectiongrowth on the tissue material becomes
Tissue-Based Membrane Electrodes of 2 x 10-5M. The response time of the visible. However, the addition of sodium
electrode is 5 to 7 minutes. The kidney azide at a concentration of 0.02 percent
The first (13) bioselective membrane tissue electrode yields negligible re- completely eliminates this problem, and
electrode made with intact animal tissue sponse to such possible interferences as electrode lifetimes of at least 30 days are
slices utilized a rather crude arrange- urea, L-alanine, L-arginine, L-histidine, routinely obtained with no significant
changes in the response characteristics
of the electrode system during that peri-
Summary. The use of intact bacterial cells or tissue slices of plant and animal origin od.
as immobilized biocatalysts has extended the possible range of potentiometric A study (15) of glutamine-selective po-
bioselective membrane electrodes beyond that of conventional enzyme electrodes. tentiometric membrane electrodes that
The use of such materials as biocatalysts offers advantages in situations where compared the use of isolated enzyme,
isolated enzymes are not available or where multistep reaction paths are required. bacteria, mitochondria, and animal tis-
The resulting bioselective electrodes also offer exceptional ease of preparation, time sue as biocatalysts clearly identified the
stability, and low cost. advantages of the tissue type in terms of
low cost, superior mechanical and time
stability, and ease of preparation. Such
ment (Fig. 1). This electrode required L-valine, L-serine, L-glutamic acid, L- electrodes may become particularly at-
both beef liver tissue and isolated urease asparagine, L-aspartic acid, D-alanine, D- tractive in situations where isolated, pu-
enzyme to mediate the conversion of the aspartic acid, glycine, or creatinine. rified enzymes are not readily available,
amino acid to be measured, arginine, to It might be expected that such animal are unstable, too costly, or require co-
the electroactive product, ammonia, via tissue electrodes would have rather short factors for proper biocatalytic activity.
the scheme: In developing nations, moreover, fresh
animal tissues are likely to be more
arginine nien)+ue
liver tissue ornithine+ urea (1)
readily available than pure enzyme prep-
urease 2 + 2NH arations.
(2)
An interesting variation on the poten-
Despite its limited utility, this early elec- tiometric tissue-based electrode concept
trode effectively demonstrates the con- has been reported by Updike and Trei-
cept of using intact tissue slices as bio- chel (16). These authors constructed an
catalysts and, moreover, illustrates the electrode with response to antidiuretic
"building-block" approach to electrode hormone (ADH) by stretching a toad
design. This approach employs a combi- bladder tissue membrane over the sur-
nation of biocatalytical components and face of a sodium ion-sensing glass elec-
membranes to give substrate selectivity trode; this arrangement permits monitor-
and to serve as physical barriers, respec- ing of potentials resulting from the en-
tively, in a manner designed to yield hanced transport of sodium ions across
overall selectivity along with electrode the toad bladder membrane in the pres-
stability and good response characteris- ence of ADH, with a rough proportional-
tics. ity between the magnitude of the poten-
The full power of this approach has tial and the hormone concentration over
been realized (14) in the design of the a narrow range. Since ADH-enhanced
glutamine-selective electrode (Fig. 2). sodium ion transport occurs in only one
The need for an auxiliary enzyme is direction-from the mucosal to the sero-
eliminated because the complete biocat- sal side of the toad bladder-the mem-
alytic functions are being carried out by f4 I brane can be oriented with the mucosal
a tissue slice from the cortex portion of b * side touching the sodium-sensing glass
porcine kidney. A slice of kidney, 0.5 electrode so that a decrease in sodium
millimeter thick, is held at the surface of ion activity is produced at the inner
b -%\\x\\\\xx\\\V\ \Wx\\ .-> c
an ammonia gas-sensing membrane elec- sensor when ADH is present in the sam-
trode by means of a dialysis membrane ple. This decrease is produced within
or a mesh of monofilament nylon. The just 10 seconds; as a result, a rapid assay
Fig. 1. Schematic of liver tissue-enzyme elec-
dialysis membrane serves to block out trode for arginine: a, dialysis membrane; b, for ADH might be feasible. Unfortunate-
high molecular weight materials, and the bovine liver tissue slice; c, urease enzyme ly, sodium transport in the toad bladder
hydrophobic gas-permeable membrane suspension; d, gas-permeable membrane; e, is altered by other hormones such as
internal electrolyte; f, combination pH elec- aldosterone, thyroxine, and angiotensin,
prevents the entry of ions into the in-
trode; g, plastic electrode body; Dl, arginine
ternal electrode elements. The pork kid- as well as by adenosine 3',5'-monophos-
substrate; 0, urea intermediate; 0, ammonia
neys are obtained from freshly killed product. (Components d to g constitute the phate and potassium or calcium ions.
animals but can be stored under refrig-ammonia gas-sensing electrode.) This lack of discrimination in membrane

288 SCIENCE, VOL. 214

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action and the interference problems in- tentiometric bacterial membrane elec-
herent to the sodium-sensing glass elec- trodes is exceptionally simple. All that is
-e
trode that serves as the inner component A I necessary is to hold a layer of bacterial
probably mean that this particular tissue cells at the surface of an ion- or gas-
electrode cannot be made selective and sensing potentiometric membrane elec-
thus will have limited practical useful- trode selected on the basis of its re-
ness. sponse to some product of the bacterial
In some cases, it becomes possible to metabolism. Although most of the suc-
improve the selectivity of a tissue-based _0 o-0- a cessful bacterial electrodes described to
electrode by "tuning" of the biochemi- date have used a selected cultured strain
Fig. 2. Expanded view of porcine kidney
cal steps involved. This was recently tissue sensor for glutamine: a, nylon mesh or of bacteria, even this is not an absolute
demonstrated (17) by the selectivity en- dialysis membrane; b, kidney tissue slice; c, requirement, as has been demonstrated
hancement of an adenosine-sensing elec- dialysis membrane; d, gas-permeable mem- by the limited but workable sugar sensor
brane; e, internal electrolyte; f, combination
trode in which mouse small intestine that uses crude human dental plaque (23)
pH electrode; g, plastic electrode body; 0,
mucosal tissue serves as the biocatalyt- glutamine substrate; *, ammonia product. spread on a pH glass electrode.
ic layer. In its untreated state, such tis- A more typical arrangement is that
sue does not effectively discriminate be- shown in Fig. 4 for the highly selective
tween adenosine and the related phos- drawn from plant tissues may, in the glutamine electrode (24). Bacterial cells
phate nucleotides, owing to alkaline future, greatly extend the possible range of the pure strain Sarcina flava (Ameri-
phosphatase enzyme activity in the tis- of tissue-based potentiometric mem- can Type Culture Collection No. 147) are
sue along with the desired adenosine brane electrodes. held at the surface of an ammonia gas-
deaminase activity. However, studies The cost of tissue electrodes is being sensing membrane electrode by means of
have shown (17) that the interfering ac-reduced through the use of very inexpen- a dialysis membrane. Although the vol-
tivity can be effectively suppressed with sive internal electrode elements (19) to a ume of cells held at the electrode surface
glycerol phosphate or by an inhibitor point where the possibility of completely is only 10 to 15 microliters, even this
such as L-phenylalanine to yield a final disposable electrodes may become a re- quantity corresponds to 108 to 109 living
electrode system that is highly selective ality. bacteria. The bacterial cell layer is ob-
to the primary substrate, adenosine. tained quite readily by culturing the ap-
Tissue-based membrane electrodes propriate strain under sterile conditions,
are still in such an early stage of develop-Bacterial Electrodes harvesting and washing the cells, com-
ment that numerous practical and funda- pacting the cells from suspension by
mental questions remain to be answered. Although Divies (20) used bacteria gentle centrifugation, and spreading the
What, for example, is the optimumwith electrochemical detection for etha-
thick- cells manually on the surface of the inner
ness of the tissue slice to be used? Ex- nol analysis as early as 1975, the first potentiometric electrode. The Sarcina
periments in our laboratory have shown potentiometric membrane electrode with flava strain is sufficiently selective in its
that the answer involves a compromise living bacterial cells serving as biocata- biocatalytic activity that this electrode
between maintaining mechanical integri- lysts appears to be the arginine-selective can be used for the measurement of L-
ty and holding dynamic response times electrode described (21) in 1976. An ex- glutamine in the presence of the other
of the electrodes to reasonable limits. In cellent review of the state of the art to essential amino acids, even in such com-
the case of a rabbit muscle tissue-based 1979 was recently given by Kobos (22). plex samples as human blood serum.
electrode for adenosine 5'-monophos- The experimental arrangement for po- Provided that care is taken to prevent
phate (AMP), we found that a tissue
thickness of 0.5 mm gives overall elec-
trode response times of 3 minutes in the
millimolar AMP concentration range; yet
even this small quantity of tissue con-
tains 50 times the effective biocatalytic
activity of an equivalent volume of the i
Fig. 3. Pictorial diagram of a
purified commercially available AMP de- i
plant tissue-based mem-
aminase enzyme preparation. brane electrode for glutamic h
The latest advance in the development acid; a, yellow squash plant;
of tissue-based electrodes is the discov- b, pericarp skin layer; c, me-
ery that materials of plant origin can socarp
be skin layer; d, endo-
carp skin layer; e, squash
used as effective biocatalysts (18). Fig- g
tissue slice from mesocarp
ure 3, for example, shows a plant tissue layer; f, immobilizing ma-
electrode devised from the mesocarp trix; g, gas-permeable mem- b
skin layer of the growing portion of a brane; h, spacer; i, internal c -:..'
yellow squash, with a carbon dioxide electrolyte; j, combination
pH electrode; k, plastic elec- d --~-
gas-sensing probe as the inner element. trode body. (Components g
The squash tissue slice serves as a bio- to k constitute the carbon
catalyst for the breakdown of glutamic dioxide gas-sensing elec-
trode.)
acid to yield products including carbon
dioxide,whose production gives rise to a
potential reading related to the concen-
tration of glutamic acid in the sample.
The possibility of using biocatalysts a

16 OCTOBER 1981 289

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contamination, useful electrode lifetimes A much more elegant solution to the
of 20 days or more are easily attained multistep sequence problem is to utilize
with no significant changes in response bacterial strains that already contain all
characteristics. Walters et al. (25) re- the necessary enzymes and cofactors to
cently improved the physical design of carry out the entire complex reaction
bacterial electrodes by substituting a f -e sequence. An exceptionally clear exam-
sterilizing filter membrane for the dialy- ple of this approach was recently report-
sis membrane in their preparation of a L- ed by Kobos and Pyon (30), who devel-
histidine-sensing electrode based on the oped a bacterial sensor for nitrilotriace-
strain Pseudomonas sp. (American Type d C tic acid. In this case, the bacterial cells of
Culture Collection No. 112996). the Pseudomonas strain must carry out a
A distinctive feature of bacterial mem- four-step reaction sequence to convert
brane electrodes is the possibility that b a the nitrilotriacetic acid to ammonia. The
their lifetimes may be extended through greatest biocatalytic activity was ob-
regeneration, that is, through the re- tained with bacterial cells harvested ear-
growth of fresh cells on the electrode ly in the exponential growth phase; as
surface. This effect has been demonstrat- a result, good sensitivity and response
ed with an electrode for L-aspartate (26), slope could be achieved, although sever-
based on the strain Bacterium cadaveris al significant interferences may limit the
at an ammonia gas-sensing membrane practical usefulness of this sensor.
e I
electrode, and the L-cysteine (27) elec- [] ?] <-.- d
trode, which uses Proteus morganii in
conjunction with a hydrogen sulfide gas- Future Prospects
sensing membrane electrode. In both
cases, the biocatalytic activity can be Fig.U-- - 4 C- .a Recent work in Suzuki's laboratory on
regenerated by placing the spent elec- the use of immobilized whole cells in
trode back into the nutrient growth medi-Fig. 4. Cross-sectional view of bacterial
lec- conjunction with ael
pH electrode in a flow
um used for the culturing of the respec- trode for glutamine: a, dialysis ; b, system membrane
(31, 32), could easily lead to new
bacterial layer; c, gas-permeable membra
tive bacterial strain. Apparently, fresh
d, internal electrolyte; e, combination
; bioselective membrane electrodes if the
cells are grown in situ at the electrode electrode;f, plastic electrode body; D, uta-
gli cells were held directly at the electrode
surface so that the initial biocatalytic mine substrate; *, ammonia product. surface. Specifically, it was shown that
activity can be largely restored and the cephalosporin (31) could be determined
electrode lifetime extended; this method with immobilized Citrobacter freundii
becomes self-limiting after a few re- method for the joint immobilization I of cells and nicotinic acid (32) with immobi-
growth cycles owing to the buildup of these several materials, and this rep pre- lized Lactobacillus arabinosus, on the
cellular debris at the electrode surface. sents a desirable simplification in the basis of the pH changes produced.
Bacterial electrodes need not be re- preparation of bioselective membr ane Work in progress in our laboratory
stricted to sensing biochemical sub- electrodes. suggests that yet a new class of bioelec-
stances. Kobos et al. (28), for example, Such a consideration becomes es ,pe- tive membrane probes may be possible
have developed a bacterial electrode forcially important when complex,tep multis
through the use of anaerobic bacteria.
the determination of nitrate ion. This biocatalytic pathways are necessary i to Intact cells of the type Clostridium aci-
convert the substrate to be determi] ned diurici immobilized at a potentiometric
electrode uses bacteria of the strain Azo-
tobacter vinelandii in conjunction with into a form that can be sensed by the ammonia gas sensor yield (33) an elec-
an ammonia gas-sensing internal elec- internal electrode element. Two ap- trode for L-serine with good response
trode. Nitrate is reduced to ammonia in proaches have been taken in such cas ses. characteristics. Although these bacterial
the two-step sequence First, "hybrid" electrodes have b een cells must be grown under anaerobic
constructed in which a bacterial str rain conditions, the final sensor can be effec-
NO3- + NADH-reductase
nitrate and a separate enzyme catalystint- aretively
jo employed in nondeaerated sam-
ly immobilized at the electrode surf ace ples. It may also be possible to prepare
N02- + NAD+ + H20 (3)
as, for example, in the nicotinamide a tde- electrodes for high-temperature mea-
nitrite nine dinucleotide (NAD) (29) electr< ode surement by using thermophilic bacteria.
NO2- + 3NADH -
reductase
system. In this case, the enzyme NA LD+ Both practical and fundamental stud-
nucleosidase was used with whole c ells ies need to be carried out on bacteri-
NH3 + 3NAD+ + 2H20 (4)
of Escherichia coli to provide the se- al and tissue-based potentiometric elec-
where both enzymes are contained quencein the trodes. There is as yet no general theo-
bacterial cells. Nitrate concentrations as ide retical formulation for the steady-state
low as 10-5M can be determined with a NAD+
NAD+ + +
H H20 >
NAD+ nucieosidase nicotinan
and time-dependent behavior of these
precision of 3 to 4 percent. + adenosine diphosphate-ribose (5) electrodes in terms of geometric and
This electrode illustrates another fa- kinetic parameters. A systematic investi-
vorable aspect of bacterial electrodes; in gation of the various possible immobili-
nicotinamide + H20 nicotinamide>
addition to the two enzymes, the cofac- deaminase
zation procedures and their effect on
tor NADH (nicotinamide adenine dinu- nicotinic acid + NH3 (6) electrode response is urgently needed.
cleotide, reduced) and a means for its Finally, considerably greater effort is re-
regeneration are all contained within the where the ammonia gas produced is quired to investigate means-through
bacterial cells in an optimized environ- sensed by the internal membrane el ec- blocking or inhibition of undesired meta-
ment. Thus, it is not necessary to find a trode element. bolic pathways-of improving the selec-
290 SCIENCE, VOL. 214

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tivity of whole cell bioselective mem- References and Notes 17. M. A. Arnold and G. A. Rechnitz, ibid. 53, 515
(1981).
brane electrodes. 1. G. H. Fricke, Anal. Chem. 52, 259R (1980). 18. ____ , Chem. Eng. News 59(9), 24 (1981).
2. P. Bergveld, N. F. DeRooij, J. N. Zemel, Na- 19. M. E. Meyerhoff, Anal. Lett. 13(B15), 1345
At present, significant research on ture (London) 273, 438 (1978); J. Janata, in Ion- (1980).
bacterial and tissue-based potentiomet- Selective Electrodes in Analytical Chemistry, H. 20. C. Divies, Ann. Microbiol. (Paris) 126A, 175
Freiser, Ed. (Plenum, New York, 1980), p. 107. (1975).
ric membrane electrodes is being carried 3. G. A. Rechnitz, Chem. Eng. News 53(4), 29 21. ____ , Chem. Eng. News 54(44), 23 (1976).
(1975).
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tal improvements may be categorized as

follows:
1) The development and use of spec-
trometers operating at higher magnetic
fields, in some cases with large and ver-
satile probe (sample) geometries.
2) The development of multinuclear
Recent
Recent Developments
DevelopmentsininNuclear
Nuclear spectrometers.
3) Improved spectrometer design for
Magnetic Resonance Spectroscopy Fourier transform techniques and higher
sensitivity for NMR with protons and
other nuclei.
George C. Levy and David J. Craik
4) Advances in computer capabilities.
Magnetic field strength. One of the
primary instrumental improvements has
been the use of stronger magnetic fields.
From modest but promising begin- most of the new uses of NMR have In the early 1960's it was rare for an
nings in the 1940's and 1950's, nuclear occurred. Of course, there are also many NMR spectrometer to be other than a 60-
magnetic resonance (NMR) spectrosco- applications in physics, and the future megahertz instrument, capable of ob-
py has developed into an important re- holds promise for the further expansion serving only sensitive nuclei such as
search tool (1). Early in its history, NMR of NMR spectroscopy into other disci- protons. Commercial spectrometers bf
spectroscopy was adapted from sole use plines. For example, routine medical di- the day were based on iron permanent or
by physicists, who had first discovered agnosis through NMR seems possible in electromagnets with fields of about 1.4
it, to the realm of chemists, who saw the the foreseeable future. Geological appli- tesla (14 kilogauss). In the middle 1960's
potential of the so-called chemical shift cations are also under development, electromagnets with fields of 2.3 tesla
phenomenon as a structural probe. This largely as a result of advances in the (equivalent to a resonant frequency of
first useful parameter has now been sup- technology associated with NMR of sol- 100 MHz for protons) became available,
plemented by many other experimentally id samples. We hope that this review will and they are still in extensive use. This
accessible quantities. In this article we demonstrate some of the strengths of field represents about the limiting
outline some of these new developments NMR to scientists not familiar with the strength of a conventional NMR magnet.
in NMR spectroscopy. technique and perhaps stimulate some However, since the early 1970's there
In a review of this scope it is not further new applications. has been increasing use of superconduct-
possible to cover all developments in an ing solenoid-based systems, which are
area. We have attempted to provide a capable of much higher magnetic fields.
brief overview of the advances in tech- Instrumentation
nology during the past several years to- George
George Levy
Levy
is a professor
is a professor
in the Department
in the Department
of of
gether with a discussion of some of the Chemistry and director of the NIH Biotechnology
In broad terms, most new applications
Research Resource for Multi-Nuclei NMR and Data
many new applications. We have of NMR in recent years have derived Processing at Syracuse University, Syracuse, New
stressed applications in chemistry and from parallel improvements in instru- York 13210. David Craik is a CSIRO (Australia)
postdoctoral research associate at Syracuse Univer-
biology, as these are the areas in which mentation and methods. The instrumen- sity.

SCIENCE, VOL. 214, 16 OCTOBER 1981 0036-8075/81/1016-0291$01.00/0 Copyright ? 1981 AAAS 291

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