253 - Development of an Implantable Electrocatalytic Glucose Sensor

BioeZeclvocile1~zislry

and

253 - Development Glucose Sensor

Bioenergetics

5, 607-624. (1975)

of an Implantable

-

Electrocatalgtic

by U. GEBH.~RDT,G. LUFT, G. J_ RICHTER and F. VOX STG’RM Forschungslaboratorien Manuscript

der Siemens

received hlai zznd

AG..

D-Sszo

Erlan,aen,

B R.D.

1978

An electrocatalytic glucose sensor wil1 be a very useful component of an artificial beta cell in controIIing the insulin dosage during the diabetes therapy. The attempt to utilize the fuel cell principle or a stationary anode proves unavailing because of the deactivation of the electrode by reaction products or co-reactants. A potential-jump technique in which the potential shifts periodically between a measuring and a rejuvena tingphase enables us to develop a method for the reproducible determination of glucose in vitro. The method also permits to carry out measurements with sufficient accuracy even However, this requires when amino acids are present simultaneously. the electrode to be covered by a membrane in order to limit the mass transfer rate of the reactants and of the co-reactants. Under the present conditions of measurement, the glucose response deviates at most by 15 oh if the amino acid concentration raises from o to the physiological masimum value.

Introduction

.

Diabetes mellitus is a disease due to metabolic disturbance. It is based on an irregular or insufficient release of insulin,by the beta cells causing an excessive sugar level in the diseased person. T About 1 oh of the population qf developed nations suffer from diabetes, but only half of them have been diagnosed and are under medical control. Nearly I o/o of the population is dependent on the consistent injection of insulin. The adaptation of the insulin concentration in blood to the rapidly changing body requirements is a long way from being perfect _ The patients must endure considerable restraints and inHarmful consequences cannot conveniences (e.g. diet and injections). be ruled out even with a careful treatment. Accelerated arteriosclerosis and organ defects can lead to high blood pressure, renal insufficiency, sight deterioration to the point of blindness and also to the 10~s of limbs by amputation.

60s

Gebhardt. Luft. Richter and van Sturm

For some time, efforts have been under progress to compensate this hormonal malfunction by an open1 or closed loop deviceZB3 for supply of insulin. Such a system is called artificial pancreas or artificial beta cell. The closed loop system consists of a glucose sensor, an insulin depot, Until now, a dosage unit, an energ>- source and an electronic device. such systems have taken the form of portable or bed-side appliances_ They sewe for emergency treatments, e.g. coma therapy- or for treatment during surgery. An implantable system is aimecl which should not be larger than a golf ball or the size of a current heart pacemaker in its design concept and Which relieves the diabetic patients from the necessityof frequent injections and possibly- minimizes or even eliminates, the subsequent organ defects by dosing insulin exactly. The bed-side implements have so far been coupled to an automatic analvzer of TECHSICOS Inc. (autoanalyzer), which works on the colorimet& principle. Recently, enzyme sensors have also been used instead. The_\- are based on the enzymatic osidation of glucose to gluconic acid in the presence of glucose osidase according to the equation : GIucose

+ 0,

+ Gluconic

acid + H@,

The glucose osidase is immobilized in a membrane which covers a platinum electrode. This electrode measures either the consumption of oxygen during the reaction or the formation of hydrogen peroxide by e’lectrochemical means.V Such a sensor is not suitable for long-term implantation because of the gradual inactivation of enzymes under body conditions. \Ve have therefore started developing an electrocatalytic glucose sensor whose life or service period is not, in theory, limited. The performance of this sensor is based on fuel cell principies.8:’ Glucose and oxygen are available only as a misture in body tissue or blood, an appropriate design of the electrochemical cell would allow their selective conversion at the electrodes.S-lo The performance of such fuel cells has been verified in laboratory esperimentslL and also established in long-term animal tests. I2 It is evident that the efficiency- of such a cell with specified dimensions of electrodes and membranes would depend on the glucose concentration in the surrounding solution_ However, the fuel cell cannot find direct application as a practically useful glucose sensor. X deactivation of the anode catalyst, especiallyin the presence of interfering endogenous substances such as urea and amino acids, leads to very low currents or calbi for the use of Iarge amounts of catalysts in a relativeIy voluminous and reaction processes would layer. 1 1 In the latter case the diffusion become slow and cause an intollerable time constant. The sensor in such a case would respond with great dela>- to a concentration change. _A few years ago, SOELDSER and Co-workers had alreadv introduced a sensor which consisted of a modified fuel cell with an additional auxThey reported iliary electrode for rejul-enating the working electrode. on animal tests with monkeys in which glucose concentration changes

Implantable

Electrocatalytic

Glucose

Sensor

609

were recorded even after an implanted period of 117 days. However, the response signal had dropped from the d range to the nA range during this period. Thus the sensor was still unsuitable for an absolute measurement of glucose, which is a prerequisite for a closed loop .a@ificial pancreas.r3 Experimental

The current-time curves were recorded with the usual potentiostatic set-up using a silverlsilver chloride or a hydrogen electrode as the reference electrode_ The potentials are always referred to the hydrogen potential in the same solution_ The potentiodynamic curves were recorded similarly by conventional techniques by overlapping the signal of a voltage sweep generator at the control voltage source of a potentiostat and tracing with on x-y recorder_ Smooth or platinized platinum was used as the working electrode.

Fig.

I.

*.

Basic circmt dia,sram of the setup for meisurement I. Potentlostat. 1 Integrator and 3_ Time-pro,g-amme controI_

In the potential-jump method, the electrode-potential was periodically reversed between the working and the rejuvenating potentials. The basic circuit diagram of our setup is represented in Fig. I_ The working and the rejuvenating potentials of the working electrode (WIZ) are preset or fked by means of two control voltage sources E, and E,

610

Gebhardt,

Luft,

Richter

and

L-OQ Sturm

respectively and reversed with the help of a time programme transmitter (3)_ The potentiostat (I) regulates the electrode potential against the reference electrode (RE) in accordance with the preset control \-oltage. The current flowing through the counter electrode (CE) and the integral of this current are registered by a recorder. The potential programme is schematicall; represented below in Fig. a. The current z’s_ time cun-c is depicted abo\-e by the dashed lines while the full-line curve >-ields the inregral during the period of measurement_

F1g

2.

3Icasunng programme and rcsponse agnal of the glucose sensor Bottom potential programme. Top current-time cur\ es (dashed lmes) and current Integral dunng the measunng penod (dra\\ n line).

After rel-erting to working potentia1, the integrator (2) is switched with deIa\- in order to separate the capacitive currents which emerge due to charge rel-ersal of the electrochemical double la>-er and from the reduction of pIatinum oxide. _A constant, o\-erlapped charge eschange which, for instance, results from the reduction of osygen in the elcctrol)-te is compensated. Elecfrodes Smooth platinum, platinized platinum and activated platinum from a Pt-Xi sheet were used as the eIectrodes in the experiments. The area of the smooth platinum electrode was 0.5 cm?. Before being used it was ground freshl!- on a polishin g wheel with a I pm polishing paste. The area of the pIatinized pIatinum was also 0.5 cm’ or o,o3 cm’. The smooth platinum surface was platinked bJ--cathodic deposition of platinum from a 3 -_5 o/o solution of hesachioroplatimc (IV) acid at a current densit) of 30 mX/cm’ for =j minutes. Platinum was thus deposited as platinum black. The electrodes were used in freshly platinized condition and also in a subsequently rejuvenated state after aging, i.e. the electrodes were not polished and re-platinized after each measurement but the surfaces were cleaned b!- polarizing for a short while at 1600 mV (RHE) and thus oxidizing the impurities.

Implantabk

Eiectrocatalytlc

Glucose

Sensor

611

The RWEEY platinum catalyst was prepared from a homogeneous alloy of Pt-Ni in the atomic proportion of I : 6-l* The activation was conducted by anodic oxidation of nickel at So OC in I M H,SO, in a potential range between 300 mV and 700 mV until the current dropped to zero. The geometric area of the electrode was 0.2 cm”. The roughness area in the factor was estimated to be 3.5 XIOI and the BET-surface active layer ranged between 30 and 36 m’/g. ELectrolyte Most of the measurements were conducted in phosphate buffer However, solution at pH 7 for the sake of experimental convenience_ the relevancy was additionally confirmed by measurement in TYRODE solution. This physiological solution, originally prepared by TYRODE as a culture medium, is composed of 137 rnM NaC1, 2.6s mM KCI, 12 mM and 12 mM NaHCO,. In CaCl,, I.05 mM NgCl,, 0417 mM NaH,PO, order to define the oxygen partial pressure during the esperiments, the electrolyte was saturated with air or argon. In the case of the T\-RODE solution, a misture of air (95 “/b) and CO, (5 %) was used to maintain the pH value constant at 7. Rengerlfs The solutions were prepared with PAZ. grade reagents in double distilled water. The amino acids were of purest grade. The amino acid mixture (xAM) contained the most common amino acids. The composition is given in Table I. Table

I.

Amino

acid

mixture

(A_~lI).

I Ammo

acid

Alanlne _Arginine Glyclne Hlstldlne Isoleucme Leucme Lysme Phenylalanme Threonme Tyrosme Valine Total

sum

I

I 1

-Ph~-slol. concn. mg/ioo cm3

o-99 2 03 I.2 4 1.10

r-70 I.30

i

o-99 1.63 I 05 Z?36

,

612

Gebhardt, Luft. Richter and van Sturrn

Solutions containing glucose or amino acids form excellent culture Therefore, they could easil) media for microbiaI growth and activity. Hence the x-alidit)- of long-term esperibe affected and get decomposed. ments will be considerabIy limited. In our experiments, the glass \-essels and the eIectrode supporter made of polyprop_ylene were steam autoclaved at 120 OC under a pressure of 2.1 bar for IO mmutes. Thus, it was possible to maintain steriIe conditions in the measurements extending up to S da_\-s.

In all short-term experiments using phosphate buffer or TTRODE solution as the electrolyte, platinized platinum electrode yielded a response signal which was dependent on the glucose concentration. Fig_ 3 illustrates the potentiodynamic cult-es wth a rotating pIatinized as an esample, platinum electrode in the presence of o, 0.1 and o 3 y/o glucose in phosphate buffer solution_

Potentlodynnmic current-denslt_\ irS potential cur\ es 1sIth a rctatlng platinized p1at1ltum electrode (area = 0 II? cm=. rotation 111= 30 s-1 and sweep-rate t’ = I \- mln-1) in the presence of o (curt-e I), o I (cun~ 2) and o 3 “/:, (cunc 3) glucose in phosphate buffer solutlon under argon atmosphere.

Sf end+s

fnfe iit e&hod

Basicall)-, a decay- of glucose osidation currents is observed in long-term potentiostatic measurements. The aging of the electrode b> recqrstallization of platinum black crystallites and aLso the blockage of the catall-st surface bJ- the inert intermediate products pIa!- a role dunng this current decay-. The sensitivity decreases at the same time.11 The electrode actlvit!can be regenerated by potcntiostatic or potentiod>-namic ano:lic oxidation. Some endogenous substances block the elctrode surfaces more strongI)- than the intermediate reaction products of glucose-I1 The strongest

Irnplantable

Electrocatalytic

Glucose

Sensor

613

inhibiting effects are created by the presence of amino acids in physiolTherefore, they served in our present work as logical concentrations. models for all other interfering endogenous substanccs.15Sr6 \Vhen a platimzed platinum electrode was polarized for a longer period in the presence of amino acids, a measurable current response as a function of glucose concentration change w-as obtained only above a potential of 1100 mV. Fig. 4 shows the results of experiments in which the electrode was potentiostated for 16 hours rn an electrolyte contaming amino acids before glucose was added at a concentration of 0.1 To. The composition of the amino acid mixture (AMI) solution is given in Table I.

Potentiostatx current-density us time cuncs wth a platmized platmum electrode m phosphate buffer solution containmg ammo acid mlsture (XXMJ under the addltlon of o I ‘J/, glucose after 16 hours, Ar-atmosphere I TOO mV ; 1100 mV; 4 2. go0 mV ; 3 1300 rn\, and 5 _ 1500 m\ ifr

t(h)

15

2’3

For the sake of simplicity, only those amino acids ha\-e been considered Even with which appear in high concentrations in plasma solution. glucose addition measurable current changes were not found up to a potential of 1100 mV. A distinct response is obtained only at 1500 ml’. The drastic influence of amino -acids is also eshibited in slow potentiodynamic curws. The quasi-stationary current density--potential curves are shown in Fig. 3 in which only the anodic part is considered. The phosphate buffer solution with glucose content (curve I) shows distinctl! two maxima for glucose osidation. But these masima disappear completely in the presence of amino acids under phy-siological concentratrons (cur\-e 3) _ Escept at very- high potentials, the presence or absence of glucose made no drfference when the electrolyte contained amino acids (curves z and 3). A comparison with the blank electrolyte (cun-e 4) shows that the osidation of amino acids begins above IOOO mV. These results lead to the conclusion that the measurement of glucose concentration should be possible if the access of glucose-amino acid mixture at the electrode is limited by a membrane to such an extent that the arriving amino acids are totally or significantly oxidized. X very active electrode and a relatively impermeable membrane will be required

Gebhardt,

61-t

Luft,

Richter

and

VOR Sturm

Quasi-stationary potentiodynamic current-density vs potential curves ~itll a platimzcd platmum electrode (area, o 2s cmz), sweeprate u zoo rnL_/h. argon atmosphere ; phosphate buffer solutxon at pH 7 as the blank electrolyte I - BE Cth 0.1 y/o glucose ; (BE). 1: BE wth XAM and o I “16

glucose ; 3 . BE 111th _A.AJL and 4 : BE alone

for this purpose. In this connection, the R..xsEs-platinum catalyst electrode had pro\-cd suitable with a catalJ-tic la)-er thickness of IOO pm and Perrnion 1035 of the RAI Research Corporation, appeared to be useful as the membrane. Fig. 6 shows the current-time curve at a potential of 1350 mV with alternatin, m 3 4ucose concentration in the electrolyte. Accordingly-, this electrode/membrane system thoroughly responds to the glucose concentration changes even in long-term tests. But the time constants become too large (in the order of hours) to be of practical USC. The reaction rate in the active layer will not be suf-

,

30 Fig.

t(h)

, 40

6.

Current-density z’s time cures wth a RasrzY-platinum electrode (prepared by activatmg Pt.-Xl alloy sheet) in phosphate buffer solution at 1350 mV with different quantitles of glucose and _%_I31as the additives Diffusion hmltatlon by a Permian xozg membrane, electrode area, o ZS cm’. actlvc la>er thickness. IOO pm.

ImplantabIe

Electrocatalytic

Glucose

Sensor

-

615

ficient. Memory effects are observed and the reaction becomes very lethargic especially after the addition of ammo acids. The tune constant of the sensor is not determined by the time lag of the diffusion through This time lag XIS determined separately by measuring the membrane. the diffusion coefficient of glucose at the membrane_ The time lag was The use of a less permeable membrane is prohibifound to be IO minutes. tive anyhow because the membrane alone would make the time constants A further acceleration of the anodic intolerably large for application. o_xidation reactions by increasing the potentials is not possible because of the accumulation of gas bubbles behind the membrane (O,-evoIution). According to these results the development of either an active current generating glucose sensor based on the fuel cell principle or a passive sensor operating under stationary condition seems to have no chance.

Glucose measurement amino acids

in

the

presence

of

constant

concentrationsof

The application of non-steady potentiodynamic orpotential-jump The method has decisive advantages over the steady-state method. continual rejuvenation of the electrode warrants at thesame time high currents, fast reaction rates and high sensitivities. The possibility of separating the measurement from the rejuvenation increases the seiecThese facts are esemplified in Fig. 7 by the poti\-ity of the reaction. tentiodynamic curves which were recorded after rejuvenating for 2

Potentlodynamic current-density vs. potential curves 117th a platmwxl platinum electrode after reJuvenating for 2 mm at 1.5 V, sleep-rate 3 V mm-l ; phosphate buffer solution as the blank electrolyte (BE). I : BE alone. I? . BE with AA&C ; 3 : BE with o I yO glucose and X_L\I and 4 : BE \mth 0.1 “/0 giocose.

n I

100

200

3a.l

!

1

440

Implantablc

Electracatalytic

Glucose

Sensor

6x7

favorable if the measurement is conducted m the presence of an amino The sensitlvlty is lower and the accuracy ‘is not satiskc1 mi.xture factory X better dlscnmmatlon is achieved at a more positive rejuvcnation potcntlal of If500 mV Fig. 9 represents the calibration kurve. The readmgs are traced through a compensated parabola. The calibratlon curve 1s more strongly bent m the cast of TYRODE solution as the clcctrolyte, especially at high @ucose concentrations It has been thus demonstrated that the detennmatlon of glucose concentration 111 phy\lologlcal mechum is possible m the presence of constant ammo acrd conccntratlons Glucose measurement in the presence of fluctuatmg concentrations of dmmo acids In order to ensure the rcahable determination of glucose even in the prcwnce of fluctuating concentratlonq of amino acids, their influence on the anodlc osldatlon of glucose should be completely or IargkIy elimiwltcd The blockmg of the electrode surface can bc prevented and the cat,llyst cdn bc mnintainccl active for glucose oxidation if a diffusioh limiting current of ‘imino CLCH~SIS estabhshed This Implies that at

Cnllbratlon curve for the dctermlnntlon of glucose in pllosphatc buffer wlutlon In the prtsjence of dm~no acids Compcnsxttd Darntwls E\pcnmental condltlonh, pl,atlnlzed plntlnum cltxtrodc ~tlth a geometric krt_, oE membrxw, 0 03 cm’, u lthout a cokenng re~u\cn~t~on at 1600 mV for 25 s snd mea\urement at 400 mV for 25 5, argon atmosphere Dela~cul s\\itch~ng of the Integ’;ltor after Y s

500 -

-%I 9.

01’ 0

Glucose %

01

CL2

03

I

I34

I

05

least dunng the reluk-cnating phase a diffusion limited current is estabz lashed for the osiclatlon of ammo acids when a suitable covenng membrane 1s used The osldatlon current is potential dependent as depicted in Fig.- 4 In view of the relatively large numbef of iamino and 3 respcctivcly acids which differ in their size, structure and concentration it is dlfficklt to make a quantitative statement regarding their diffusion through’ the

Ccbhardt,

GrS

Luft,

Richter

and

von

Sturm

mcmbrancs. A qualitative comparison is illustrated in Table 2. It contains diffusion cocfficicnts of glucose for various membranes (column 3). The glucose molccuk is comparable in its size with that of amino acids. The flow of diffusion is calculated as the diffusion limiting current under the assumption of complete oxidation of glucose (24 electrons) and a concentration of 0.1 o/o (column 4). I-~i,lC 2.

Diffusion

of glucose

-I-

through

membranes_

I

2

3 D

d

Cl11 _ _e----e----:

!

cm2 s-l _-- _.I_

-_

I

I i -I-

4

illm*

A

cm-2

I

5

=** /

S

1 1

1-S

1.0

s 10-z

3 3 x

_I 5 s x0-3

2 4

I ss

I

x0-3

x 10-e

x

10-S

x 10-d

q-2

x 10-5

77 506

10-i

14

ssxo-’

_I 5 s 10-S

11x10-6

13s IO-) I -2 x x0-1 2-j s 10-J

10-T s-7 s 10-S 5

2.5

ss

2.3 s xc-s

3

3-I s 10-S

1 ]

3 15’ 276

3541

The ditt‘usion Limitation in the case of a relatively slow reaction, such a~ the oxidation of amino acids, can be achie\-ed by increasing the potcncinl or b>- incrensin,= the electrode acti\-it\-, ~.g., by strongly roughwing tht> elcctrock surface. An improl-cd act;\-ity howe\-er, would lead in the c.~c of the potential-jump method to an unwanted rise in energ>consumption due to the neccssar>- capaciti\-e charge eschange. Finall)-, the transport limitation through a compact less permeable membrane would also remain yre\--alent_ The application of a less permeable memhrxn~ Iead~ at the SUIW time to a longer time constant in detecting the conxntnrion chnngcs of the solution at the electrode surface_ The time const‘\nt 2, which is =also referred as time lag is calculated according to JOST’” from the expression

Implantable Electrocatalytic Glucose Sensor 3d* In 3 2X” D

Z,=--.-

.

619

d” = 0.167 D

The time constants of various membranes have been calculated and given in the last column of Table 2. In considering a controlled dosage of insulin by means of an artificial beta cell, it is essential that the time constant of the sensor should not be longer than that of the glucose level adjustment inside the body (approximately 7 min.). The last membrane in the Table (Permion 5010) does not fulfill this requirement. If we succeed in achieving a diffusion limitation for the anodic oxidation of amino acids, their influence on glucose oxidation should remain low and uncritical. Although the total concentration of all amino acids amounted to 0.05 %, which is half the value of glucose used, most of the amino acids are more diEcult to oxidize than glucose in the working potential range of 300 - 400 mV. A comparison of the current-time curves at the working potentials in Fig. IO and II respectively shows the simultaneous effect of the diffusion inhibition through a membrane and of the rejuvenating potential.

Fig. IO. .

Current density vs. time curv es at the working potential of 300 mV after a rejuvenation for 15 s at IZOO mV. electrode area o 03 cm’, argon-saturated phosphate buffer solution as the blank electrolyte (BE) _ I - BE alone ; 2 : BE wqth XAM ; 3 _ BE wth o I :4, glucose and d.Ul ; _I a BE with 0.x o/O glucose. - *

The curves emerge from the negative region, below left, because, after the potential reversal from the rejuvenating potential to the working potential, the reduction of the surface oxide layer occurs at the platinum electrode. Curve I shows the respective behavior of the blank electrolyte (BE) - phosphate buffer or TYRODE solution. Curve 2 corresponds to the electrolyte containing amino acid misture. Curve 3 belongs to the electrolyte solution containing amino acid mixture and glucose, whereas curve 4 depicts the blank electrolyte containing only glucose and without amino acids.

Gebhardt,

Luft,

Richter

and

van

Sturm

Fig_ II. Current density DS.tlmc cun es at a \\orklng potential of +X-J mV after a rqukenation for 25 s at 1600 mv. electrode area o 03 cm’. co~cred with a membrane (SSRTORIUS I 1739). electrol! te ; TYRODE solution saturated 117th a-gas mwture of air (95 y&) and co, (5 0’ ,J - Sigmficance of the curves r-4 is the same as m the preceding figure (Fig IO)

0

Go2 go1

t

I

3

1

if-c

150 z

2

ty_

IOO, 50 0

I

100

I

200

3

n 300

1 400

Influence of amino acid concentration on the rcsponse of a glucose sensor, ~a= number of measuring qcles, electrode area o 03 cm=. covered wth Perrmon 102s membrane ReJuvenatxon at I 600 m\- for rj s and measurement at 400 mV for 2s s. delay time of the integrator I 5 s Electrolyte scavenged with a gas mwture of air (95 %) and 101% est ammo acid concentrahon co, (5 Yb). I (column 2, Table 3) : 2 - lughest ammo acid concentration (column 3. Table 3) ; 3 : one-half of the highest ammo acid concentration

Implantable

Electrocatalytic

Glucose

Sensor

621

The response signal (J j dt) is referred to the blank electrolyte free of glucose but with or without amino acids. This is characterized by the area between the curves I S- .+ and a & 3 respectively. Fig. IO shows the drastic effect of amino acids on the glucose reaction. In this case the electrode was uncovered and reluvenated at IZOO mV. Fig. II shows however, that the influence of amino acids can be largely suppressed by covering the electrode with a suitable membrane and rejuvenating rt at 1600 m\;. Thus, at constant levels of amino acids it is possible to overcome their adverse effect on glucose reaction mechanism_ This result is confirmed in further experiments even under fluctuating concentrations of amino acids. The concentration variations of amino acids within the ph)-siological range and their influence on the response signal during the glucose concentration measurement are illustrated in Fig. 12. The fluctuation range of amino acid concentrations has been variously reported by several authorsLs-“1 We have used the concentration range as reported by DICKIXSOS EL CZZ.~~ The lower and upper limiting values are presented in Table 3. The amino acid level in blood is considcrabl_!- raised for several hours after a protein rich meal. An elevated ammo acid le\-el is also observed dyring diabetes mellitus_22-24 Only valine and leucine were found to drop below the given fluctuation range while the total sum increases only by about IO %_ In the esperiments represented by Fig. 13, the electrode was covered with a Permion 1035 membrane, TYRODE solution served as the After the response signal became constant in a soIution electrolyte. of glucose was increased containing 0.1 o/0 glucose, the concentration The lowest concentration of amino acid was to 0.2 s/o (see upper part). added at the point marked I (Table 3, column a)_ The signal is slightly increased and it remains nearly constant at this level. It follows that neither the electrode surface is blocked nor is the glucose reaction inhibited by the presence of amino acid. The concentration of amino acid was then raised to the ma_ximum value at the point a. The renewed increment in response signal now indicates a contribution of amino acid to the total current. In this case, the total influence of amino -acid at concentration levels between o and the maximum value corresponds to a relative deviation of glucose concentration by 15 O/,_ A similar deviation (cn. 13 %) is observed when the glucose concentration is taken back to the initial value of 0.1 o/o by diluting with an electrolyte containing the maximum Reducing the amino acid concentration concentration of amino acid. to half its masimum \-alue by dilution with an electrolJ-te containing glucose lowers the deviation to 7 oh or less of the initial value (18 < 50). Even in this extremely unfavorable case, the variation of amino acid concentration from o to the maximum value would bring about a 15 o/o higher indication of glucose concentration which would be still Under the actual and physiological conditions, the amino tolerable. acid concentrations do not fluctuate from o to the maximum value but only between a minimum and masimum value. Normally with the food reception, the physiological concentration

622

Gebhardt,

Table 3.

Mmimum

and

Luft,

masimum

Richter

and

van

concentrations

Sturm of

amino

acids

in plasma

of adults_ -

_imino

acids

lowest 2

I

I-

I

I

_.lanlne Xrgimne Cysteine Cystine GIutamic acid Glycme Histidme Isoleucme Leucine Lysine 3Iethlonine PhenylaIamne Serme Threonine Tryptophan Tyrosme Valine

12

I

I

1

4-6

11s

I I

- I 2,

13 6

_

3

97

9-3 21 I 2.3 63 6.S IZ 1

/

highest

33 7 ‘7 3 36 6 14-5

2.5 10s

I i

;

mg/l (3-3_) 4

/

44 7 26 3 ‘3.5

3

56 46 1r-5

;

Difference

Concentrations in mg/l

I

I

I ; f

i I ii

1-s /-

30s

39 19 2 20 3 29 3 r-t-9

26 *I 6 3

I

of amino acid increases along with the simultaneous rise in glucose level; Therefore, a state at which insulin has got to be dispensed anyhow. the increased indication of glucose would not be critical for a sensor which forms a component part of an artificial beta cell. A certain COregulation of insulin admission by amino acids could even be desirable. However, adequate resuIts are not available to support this assumption. The described method thus permits the reliable determination of glucose under physiological conditions in the presence of amino acids. The amino acids served in our esperiments as a model for interfering intracorporal substances, particuiarly because they severely disturb the anodic oxidation of glucose. In a similar manner, we hope to bring other interfering substances under control. Urea and alcohol should be specially mentioned in this connection_ SOELDKER and Co-workers claimed that the>- could largely eliminate the influence of concentration changes of urea on glucose osidation.‘” Problems to be solved The interfering

task of ehminating the simultaneous influence substances will be attempted in the near future.

of various But abo\-c

Implantable

EIectrocataIytic

GIucose

Seusor

623

all the problems concerning the membranes will remain to be solved. The membranes should fulfill a series of requirements simuItaneously. They include : long time physical and chemical stability, and constant permeability under the conditions of chronic contact with body tissue or blood on one side and with the catalytically active and strongly oxidizing electrode on the other side, moreover they should be reproducible in preparation and be compatible and sterilizable. Also the compatibility of the reaction products has to be ascertained and ensured. In this connection, the pH changes at the working and counter electrodes have to be specially considered lest they should lead to adverse reactions. The ultimate goal of the development is to miniaturize. the sensor consisting of working, reference and counter electrodes to a single probe unit and to adapt the entire system with respect to its voIume and power Even the partial consumption to the specification of implantation. fulfihment of some of the requirements will be good enough to serve the immediate purpose of using the sensor or its control unit for external or short-term application. Acknowledgement \Ve wish to thank Miss H. I$-IESE for her valuable experimental support in measuring the membrane permeabilities. References R REXCSER. K.D. HEPP, H. MEHSERT, hI FRASETZKI, H. KRESSE and H. GEISES, Contzntrorrs Imsd~~z Therapy wzth a Portable Mznzatrcvz=ed Infusoon Systenz. European Xssociatlon for the Study of Diabetes, _hnual Meeting. Genova, 2S-30 September, 1977 TAVAS and E.F PFEIFFER. C.H. THIMXI. W_ BEISCHER, ‘S. RAPTIS, G.Y Thevapzewoche 26 (No. 11). rg$3 (1976) _A.M_ ALBISSER and B.S LEIBEL. Pvogvess

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Elzdocvzne

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irz Tovo~zto.

1% 361 (1973) J.R. Rxo and F. s. STURXI, German patent aa oo 119 - +I.xg72 S. AISESBERG and 11; 1’. CHASG. US patent 3s 37 339 - 3_z_Ig7a R.F. DRAKE, B Ii. KUSSEROU-, S NESSISGER and S MATSUDA. Tvatzs Am. SOC. Avtzf. Intevlz Ovgaazs 16, xgg (1970) SK. \i’OLFSON, Jr.. S. J. YAO, A. GEISEL and H.R. CASH. Jr., Trans. Am. Sot. Artzf. Ipztevtz. Organs 16, rg3 (1970) J_ GISER, G.L HOLLECI~, i\t TURCHAS and R FRACALX, An Implaazfable Fuel Conf_,

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(1971)

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Elzevgy

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E*zg.

624

Gebhardt,

Luft, Richter and yon Sturrn

J . R . RAo, G . J . RICHTER, F. V. STtIR.~t a n d E . ",VEIDLICH, B i o e l e c t r o c h e m . Bzoe~zerg. 3, 139 (1976) E . ~,'EIDLICH, G . R I C H T E R , F . v . STLIRM, J . R . RAO, A. THOREN a n d H . L A GERGRES, B i o m a t e r . Died. De~,ices : l r t i f . O r g a n s 4 (3-4), 2 7 7 ( I 9 7 6 ) I~.rkV. CHANG, G. AISENBERG, J . S . SOELDNER a n d J . M . H I E B E R T , T r a n s . A m . Soc. _/trtif. I n t e r n . O r g a n s 19, 332 (1973) 14 U. GEBHARDT, J . R . RAO a n d G . J . RICHTER, J . . 4 p p l . E l e c l r o c h e m . 6, 1"7 ( 1976) Is J R. RAO, G . J . RtCtITER, G. L b ' r T a n d F v. STURM, B t o m a t e r . 3 l e d . Devices .lrt~f. O r g a n s 6, 127 ( I 978) 16 J . R . GUYTO_~, K.XV. CHAXG, S. AmENBERG a n d J . S . SOELDNER, M e d . I n s l i t , re. 9, 2~- 7 (1975) l~" 13-. GEBHARDT, G LraFT, G . RrcHTER a n d F v. SruR~t, B t o n , ed. T e c h . 22, 399 (1977) is ~,V. J o s T , Diffuszo~z ; :~[elhoden der .~'less~tng ~Lnd .duswertu~zg, V e r l a g D r. D m t r i c h S t e m k o p f f , D a r m s t a d t (z957) P- 57 i9 K. DrE,~I a n d C. L E N T S E R , D o c u m e n t a Geigl,, I V z s s e n s c h a f l h c h e T a b e l l e n , 7- Auflage_ C i b ~ - G e i g . v A G . . ]Da_~el (1976) p. 57 o 20 DICKINSON et a l , P e d t a t r z c s 36, _, (1965) 21 P . SONPART, ill .,-/snzno d 6 t d P o o l s , J . [--[OLDEN ( E d i t o r ) , E l s e v i e r P u b l i s h e r s , A m s t e r d a m (I962) p. 22o ~-: J . V~VAHRE.~, 1t;). F E L I G , E CERASI a n d R L U F T , J . C l i n I n v e s t . 5 1 , t S 7 o (I972) 2:1 I f . B I C K E r , S c h ~ ' e z : . . l i e d Ilrochenschr. 9 1 , 1397 ( I 9 6 1 ) '-'~ G.A. A D I m . E . L . MORSE a n d P M A.MIN, J L a b . C h n . M e d 86, 395 (1975) J o s h n D i a b e t e s F o u n d a t i o n , I nc ., S c m n t i f i c P r o g r e s s R e p o r t , D e v e l o p m e n t o f a 3[tnzatrtre Glucose S e n s o r f o r Use w i t h a n _drt*ficzal B e l a Cell, ,_~ovember x, r 9 7 7 It 12 13