- Email: [email protected]
Flow-injection determination of meptazinol with electrochemical detection
Chimica Acta, 111 (1979) 281-285 o Elsevier Scientific Publishing Company, Amsterdam
Analytica
-
Printed in The Netherlands
Short communication
FLOW-INJECTION DETERMINATION ELECTROCHEMICAL DETECTION
OF MEE’TAZINOL
WITH
H. K. CHAN* Wyeth Laboratories, (G t. Britain)
Huntercombe
Lane
South,
Maidenhead,
Berhshire.
SL6
OPH
A. G. FOGG Department of Chemistry, Leicestershire. LEl I 3TU
(Received
Loughborough (Gt. Britain)
University
of Technology.
Loughborough,
16th May 19i9)
Summary. A flow-injection analysis system incorporating a glassy carbon voltammetric detector cell is described_ Meptazinol (0.01-10 pg ml-‘) can be determined by electrochemical osidation in a carrier stream of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol at sampling rates up to 80 samples per hour.
Previous studies [l, 21 have shown that several phenolic analgesics and two anti-inflammatory drug compounds undergo electrochemical oxidation at a glassy carbon electrode in 0.1 IM sodium acetate-O.1 M acetic acid in 98% ethanol. Although the voltammetric procedure developed for the determination of these drugs and their pharmaceutical dosage forms was simple and rapid, it lacked sensitivity for the determination of one of the phenolic analgesics, meptazinol [m-( 3ethyL1-methyl-hexahydro-lH-azepin-3-yl)phenol hydrochloride], in biological fluids where concentrations are usually at the pg ml-’ level. A gaeliquid chromatographic procedure has been described [ 31 for determination of the drug in plasma after extraction, but this requires at least 3 ml of plasma to achieve a detection limit of 30 ng ml-‘. A liquid scintillation counting procedure [4] has been used for pharmacokinetic and metabolic studies of the drug but this gives a detection limit of only 20 ng ml-‘. In view of the lack of a simple, selective and sensitive method of determining meptazinol, it was decided to investigate an adaptation of the static voltammetric method [2] to hydrodynamic voltammetry [5] by using the flow-injection technique of R%iCka and co-workers [6,7] _ A commercially available system [8] was modified; this system comprises a flow-injection block and a glassy carbon electrochemical detector based on the wall-jet principle [9, lo]_ Small volumes (5-10 r.cl) of sample solution are injected into a carrier stream of supporting electrolyte, and the anodic current is monitored as the sample passes the electrode surface and the depolarizer is oxidized at a suitable applied potential_ As most of the sample solution
282
hits the surface of the glassy carbon electrode, approximately 80% of the depolarizer is oxidized and the method is extremely sensitive: picogram amounts of an electroactive substance can be detected at detector volumes of only about 2 ~1. The present communication describes a flow injection-voltammetric procedure for the determination of meptazinol at the 100-pg level. Modifications to the manufacturer’s instructions are described which improve the reliability of the technique_ Experimental Appamtus. A commercially available Flow-Injection Stand (Metrohm E634) was used with some modification_ The electrochemical detector was housed separately in a metal container (30-cm3 cell (EA 1069/2) capacity) to cut out external signals. A hole was drilled to introduce an 80-cm length of Teflon tubing (l-5/0.3 mm) between the injection block and the detector cell to ensure a sufficient time-lag between injection of the sample and the appearance of the corresponding current peak. The glass bottle containing the supporting electrolyte was connected directly to a nitrogen cylinder which provided a constant pressure of 1 bar to give a steady flow of supporting electrolyte of 1.2 ml min-’ through the detector cell. A PAR 174A polarographic analyser (Princeton Applied Research Corp.) was used to control the applied potential and to measure the resulting current with a sensitivity of 10 V output for a 20-nA signal. A Servoscribe recorder (Model RE511.20) fitted with an attenuator (two 10 kfi resistors in series across the terminals) was used to give a full scale deflection of 10 V on the 5-V range; the chart speed was 600 mm h-i. The conventional three-electrode configuration used comprised a working glassy carbon electrode (EA 286/l, nominal surface area O-18 cm’), a platinum counter eIectrode (EA 286/2) and a silver-silver chloride reference electrode (EA 442). The reference electrolyte was 1 M lithium chloride in ethanol presaturated with silver chloride; with this electrolyte the potential for oxidation of the drug in the recommended supporting electrolyte was sufficiently reproducible over long periods of time. In order to achieve smooth baselines, the reference electrode had to be placed downstream adjacent to the effluent outlet of the detector cell. Injections were made with a lo+1 microsyringe (Scientific Glass Engineering Pty. Ltd.). Reagents_ The purity of meptazinol hydrochloride (Wyeth Laboratories) was confirmed by thin-layer chromatography_ Other reagents were of analytical-reagent grade. For the supporting electrolyte, dissolve 8.2 g of anhydrous sodium acetate in 20 ml of distilled water, add 5.8 ml of glacial acetic acid and dilute to 1 1 with absolute ethanol; filter through a Fluoropore 0.2~pm filter prior to U.W.
For the standard meptazinol hydrochIoride solution, dissolve 100 mg of meptazinol hydrochloride in 100 ml of 95% ethanol.
283
Calibration graphs. Prepare working solutions of meptazinol hydrochloride (0.01-10 ,ug ml-‘) by dilution of the 1 mg ml-’ standard with 95% ethanol. Inject lo-p1 aliquots under the instrumental conditions described below, and plot graphs of peak height (nA) versus concentration of the injected samples (ng). Procedure. Potentiostat the working electrode at + 1.2 V versus the recommended silver-silver chloride electrode for 20 min in a flowing stream of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol (1.2 ml min-‘) at a current sensitivity of 5 PA. Set the detector potential at + 1.03 V before adjusting to the required current sensitivity in the range @.05-0.5 ,uA fs.d. Inject 10 ~1 of meptazinol solution and measure the peak heights. Results
and discussion
In flow-injection analysis with this wall-jet electrochemical detector, it is important to ensure that the flow rate is constant and pulse-free_ A constant flow rate of 1.2 ml min-’ was achieved by using Teflon tubing of 0.3 mm id. at a constant nitrogen pressure of 1 bar. Adsorption occurs at the glassy carbon electrode surface after repeated injections of meptazinol solutions, and the surface was reconditioned by cycling the potential between cathodic and anodic limits until the original characteristics were restored_ The working electrode was first potentiostated at -0.2 V for 5 min at a current sensitivity of 5 PA and then at + 1.2 V for 5 min in a flowing stream of the recommended supporting electrolyte. Subsequently, the potential of the detector was set at + 1.03 V, and the peak heights obtained for meptazinol were identical to those obtained initially_ Adsorption effects were apparent after about 40 injections. A linear-sweep voltammogram for the oxidation of meptazinol(50~g’ml-‘) in a quiescent solution of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol with the recommended three-electrode system is shown in Fig. 1. The current-potential curve shown in Fig. 2 depicts a typical sigmoidal hydrodynamic voltammogram with a well-defined plateau region, analogous to classical d-c. polarography and rotating electrode voltammetry. This was constructed from the peak currents obtained by lo-111 injections of meptazinol solution (20 r.rgml-‘) into the flow stream at each applied potential. The optimum potential of + 1.03 V was determined from this curve. Calibration graphs of peak current (nA) against the amount of depolarizer (ng) in the injected samples were rectilinear over the range 0.01-10 r.lgml-’ under the conditions recommended_ The stability of the detector cell was checked; neither the potential of the reference electrode nor the performance of the working electrode changed during the course of the experiments_ This is in contrast to initial results obtained by using the manufacturer’s instructions_ It is essential that the
reference electrode be placed after the detector electrode in the flowing stream. Further, large drifts in potential occurred when the internal reference solution in the reference electrode was not presaturated with silver chloride.
284
0.9 3
600
I
/ /*
Q :
1fiA
/-
/
600
//-
i o-2
04
APPLIED
136
08
POTENTIAL
1.0 (Vr,.Ag
l.2 AgCIl
0.7 0
0.60 APPLIED
0.9 0 POTENT1
100 AL
(V
vrr.Ag
I.1 0 ABC11
Fig. l_ Typical linear-sweep vokammogram of meptazino1 (50 pg ml-‘) in 0.05 M sodium acetate-O-1 h1 acetic acid in 9S% ethanol at the gIassy carbon electrode_ Scan rate, 5 mV s-‘_ Fig. 2. Hydrodynamic current-potential stream of 0.05 hl sodium acetate-O.1 min-’ ; sample injected 10 ~1..
curve for meptazinol (20 pg ml-‘) in a flowing M acetic acid in 9S% ethanol. Flow rate 1.2 ml
I
;
_..
_
zrnln
._._.-
---
-1 SCAN
Fig. 3. Current vs. time curves for meptazinol (0.01 JL~ml-‘) in a flowing stream of 0.05 JI sodium acetate-O.1 h4 acetic acid in 98% ethanol. Potential + 1.03 V vs. Ag/Ag Cl; flow rate 1.2 ml min-’ ; current sensitivity 2.5 nA cm-‘; sample injected 10 ~1 (i.e. each peak represents 100 pg of meptazinol); chart speed 60 cm h-l.
Figure 3 shows ten IO-r.li injections of 0.01 pg ml-* meptazinol solution (Le. 100 pg per injection) at a sampling rate of up to 80 injections per hour; the coefficient of variation was 72%. The high sensitivity is due to efficient mass transfer and the fact that at constant applied potential charging current is eliminated. The application of this procedure to the determination of meptazinol in biological fluids following a simple clean-up procedure, is under study, but low concentrations require a chromatographic separation prior to electrochemical detection_
285 REFERENCES 1 H. K. Chan and A. G. Fogg, Anal. Chim. Acta., 105 (1979) 423. 2 H. K. Chan and A. G. Fogg, Anal. Chin-r. Acta, 108 (1979) 205. 3 M. T. Rosseel, M. G. Bogaert, F. M. Belpaire and W. Oosterlinck, Cur-r. Med. Res. Opin., 3 (1975) 181. 4 R. A. Franklin, A. Aldridge and C. de B. White, Br. J. Clin. Pharmac., 3 (1976) 49’7. Prentice-Hall, Englewood Cliffs, 5 V. G. Levich, Physicochemical Hydrodynamics, N-J., 1962. 6 J. RG%zka and E. H. Hansen, Anal. Chim. Acta, 7E (1975) 145. 7 J. R%%ka and J. W. B. Stewart, Anal. Chim. Acta, 79 (1975) 79. 8 P. Gilgen and P. Rach, Chimia, 32(9) (1978) 345. 9 J. Yamada and H. Matsuda, J. Electroanal. Chem., 44 (19’73) 189. 10 B. Fleet and C. J. Little, J. Chromatogr. Sci., 12 (1974) 747.
Analytica
-
Printed in The Netherlands
Short communication
FLOW-INJECTION DETERMINATION ELECTROCHEMICAL DETECTION
OF MEE’TAZINOL
WITH
H. K. CHAN* Wyeth Laboratories, (G t. Britain)
Huntercombe
Lane
South,
Maidenhead,
Berhshire.
SL6
OPH
A. G. FOGG Department of Chemistry, Leicestershire. LEl I 3TU
(Received
Loughborough (Gt. Britain)
University
of Technology.
Loughborough,
16th May 19i9)
Summary. A flow-injection analysis system incorporating a glassy carbon voltammetric detector cell is described_ Meptazinol (0.01-10 pg ml-‘) can be determined by electrochemical osidation in a carrier stream of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol at sampling rates up to 80 samples per hour.
Previous studies [l, 21 have shown that several phenolic analgesics and two anti-inflammatory drug compounds undergo electrochemical oxidation at a glassy carbon electrode in 0.1 IM sodium acetate-O.1 M acetic acid in 98% ethanol. Although the voltammetric procedure developed for the determination of these drugs and their pharmaceutical dosage forms was simple and rapid, it lacked sensitivity for the determination of one of the phenolic analgesics, meptazinol [m-( 3ethyL1-methyl-hexahydro-lH-azepin-3-yl)phenol hydrochloride], in biological fluids where concentrations are usually at the pg ml-’ level. A gaeliquid chromatographic procedure has been described [ 31 for determination of the drug in plasma after extraction, but this requires at least 3 ml of plasma to achieve a detection limit of 30 ng ml-‘. A liquid scintillation counting procedure [4] has been used for pharmacokinetic and metabolic studies of the drug but this gives a detection limit of only 20 ng ml-‘. In view of the lack of a simple, selective and sensitive method of determining meptazinol, it was decided to investigate an adaptation of the static voltammetric method [2] to hydrodynamic voltammetry [5] by using the flow-injection technique of R%iCka and co-workers [6,7] _ A commercially available system [8] was modified; this system comprises a flow-injection block and a glassy carbon electrochemical detector based on the wall-jet principle [9, lo]_ Small volumes (5-10 r.cl) of sample solution are injected into a carrier stream of supporting electrolyte, and the anodic current is monitored as the sample passes the electrode surface and the depolarizer is oxidized at a suitable applied potential_ As most of the sample solution
282
hits the surface of the glassy carbon electrode, approximately 80% of the depolarizer is oxidized and the method is extremely sensitive: picogram amounts of an electroactive substance can be detected at detector volumes of only about 2 ~1. The present communication describes a flow injection-voltammetric procedure for the determination of meptazinol at the 100-pg level. Modifications to the manufacturer’s instructions are described which improve the reliability of the technique_ Experimental Appamtus. A commercially available Flow-Injection Stand (Metrohm E634) was used with some modification_ The electrochemical detector was housed separately in a metal container (30-cm3 cell (EA 1069/2) capacity) to cut out external signals. A hole was drilled to introduce an 80-cm length of Teflon tubing (l-5/0.3 mm) between the injection block and the detector cell to ensure a sufficient time-lag between injection of the sample and the appearance of the corresponding current peak. The glass bottle containing the supporting electrolyte was connected directly to a nitrogen cylinder which provided a constant pressure of 1 bar to give a steady flow of supporting electrolyte of 1.2 ml min-’ through the detector cell. A PAR 174A polarographic analyser (Princeton Applied Research Corp.) was used to control the applied potential and to measure the resulting current with a sensitivity of 10 V output for a 20-nA signal. A Servoscribe recorder (Model RE511.20) fitted with an attenuator (two 10 kfi resistors in series across the terminals) was used to give a full scale deflection of 10 V on the 5-V range; the chart speed was 600 mm h-i. The conventional three-electrode configuration used comprised a working glassy carbon electrode (EA 286/l, nominal surface area O-18 cm’), a platinum counter eIectrode (EA 286/2) and a silver-silver chloride reference electrode (EA 442). The reference electrolyte was 1 M lithium chloride in ethanol presaturated with silver chloride; with this electrolyte the potential for oxidation of the drug in the recommended supporting electrolyte was sufficiently reproducible over long periods of time. In order to achieve smooth baselines, the reference electrode had to be placed downstream adjacent to the effluent outlet of the detector cell. Injections were made with a lo+1 microsyringe (Scientific Glass Engineering Pty. Ltd.). Reagents_ The purity of meptazinol hydrochloride (Wyeth Laboratories) was confirmed by thin-layer chromatography_ Other reagents were of analytical-reagent grade. For the supporting electrolyte, dissolve 8.2 g of anhydrous sodium acetate in 20 ml of distilled water, add 5.8 ml of glacial acetic acid and dilute to 1 1 with absolute ethanol; filter through a Fluoropore 0.2~pm filter prior to U.W.
For the standard meptazinol hydrochIoride solution, dissolve 100 mg of meptazinol hydrochloride in 100 ml of 95% ethanol.
283
Calibration graphs. Prepare working solutions of meptazinol hydrochloride (0.01-10 ,ug ml-‘) by dilution of the 1 mg ml-’ standard with 95% ethanol. Inject lo-p1 aliquots under the instrumental conditions described below, and plot graphs of peak height (nA) versus concentration of the injected samples (ng). Procedure. Potentiostat the working electrode at + 1.2 V versus the recommended silver-silver chloride electrode for 20 min in a flowing stream of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol (1.2 ml min-‘) at a current sensitivity of 5 PA. Set the detector potential at + 1.03 V before adjusting to the required current sensitivity in the range @.05-0.5 ,uA fs.d. Inject 10 ~1 of meptazinol solution and measure the peak heights. Results
and discussion
In flow-injection analysis with this wall-jet electrochemical detector, it is important to ensure that the flow rate is constant and pulse-free_ A constant flow rate of 1.2 ml min-’ was achieved by using Teflon tubing of 0.3 mm id. at a constant nitrogen pressure of 1 bar. Adsorption occurs at the glassy carbon electrode surface after repeated injections of meptazinol solutions, and the surface was reconditioned by cycling the potential between cathodic and anodic limits until the original characteristics were restored_ The working electrode was first potentiostated at -0.2 V for 5 min at a current sensitivity of 5 PA and then at + 1.2 V for 5 min in a flowing stream of the recommended supporting electrolyte. Subsequently, the potential of the detector was set at + 1.03 V, and the peak heights obtained for meptazinol were identical to those obtained initially_ Adsorption effects were apparent after about 40 injections. A linear-sweep voltammogram for the oxidation of meptazinol(50~g’ml-‘) in a quiescent solution of 0.05 M sodium acetate-O.1 M acetic acid in 98% ethanol with the recommended three-electrode system is shown in Fig. 1. The current-potential curve shown in Fig. 2 depicts a typical sigmoidal hydrodynamic voltammogram with a well-defined plateau region, analogous to classical d-c. polarography and rotating electrode voltammetry. This was constructed from the peak currents obtained by lo-111 injections of meptazinol solution (20 r.rgml-‘) into the flow stream at each applied potential. The optimum potential of + 1.03 V was determined from this curve. Calibration graphs of peak current (nA) against the amount of depolarizer (ng) in the injected samples were rectilinear over the range 0.01-10 r.lgml-’ under the conditions recommended_ The stability of the detector cell was checked; neither the potential of the reference electrode nor the performance of the working electrode changed during the course of the experiments_ This is in contrast to initial results obtained by using the manufacturer’s instructions_ It is essential that the
reference electrode be placed after the detector electrode in the flowing stream. Further, large drifts in potential occurred when the internal reference solution in the reference electrode was not presaturated with silver chloride.
284
0.9 3
600
I
/ /*
Q :
1fiA
/-
/
600
//-
i o-2
04
APPLIED
136
08
POTENTIAL
1.0 (Vr,.Ag
l.2 AgCIl
0.7 0
0.60 APPLIED
0.9 0 POTENT1
100 AL
(V
vrr.Ag
I.1 0 ABC11
Fig. l_ Typical linear-sweep vokammogram of meptazino1 (50 pg ml-‘) in 0.05 M sodium acetate-O-1 h1 acetic acid in 9S% ethanol at the gIassy carbon electrode_ Scan rate, 5 mV s-‘_ Fig. 2. Hydrodynamic current-potential stream of 0.05 hl sodium acetate-O.1 min-’ ; sample injected 10 ~1..
curve for meptazinol (20 pg ml-‘) in a flowing M acetic acid in 9S% ethanol. Flow rate 1.2 ml
I
;
_..
_
zrnln
._._.-
---
-1 SCAN
Fig. 3. Current vs. time curves for meptazinol (0.01 JL~ml-‘) in a flowing stream of 0.05 JI sodium acetate-O.1 h4 acetic acid in 98% ethanol. Potential + 1.03 V vs. Ag/Ag Cl; flow rate 1.2 ml min-’ ; current sensitivity 2.5 nA cm-‘; sample injected 10 ~1 (i.e. each peak represents 100 pg of meptazinol); chart speed 60 cm h-l.
Figure 3 shows ten IO-r.li injections of 0.01 pg ml-* meptazinol solution (Le. 100 pg per injection) at a sampling rate of up to 80 injections per hour; the coefficient of variation was 72%. The high sensitivity is due to efficient mass transfer and the fact that at constant applied potential charging current is eliminated. The application of this procedure to the determination of meptazinol in biological fluids following a simple clean-up procedure, is under study, but low concentrations require a chromatographic separation prior to electrochemical detection_
285 REFERENCES 1 H. K. Chan and A. G. Fogg, Anal. Chim. Acta., 105 (1979) 423. 2 H. K. Chan and A. G. Fogg, Anal. Chin-r. Acta, 108 (1979) 205. 3 M. T. Rosseel, M. G. Bogaert, F. M. Belpaire and W. Oosterlinck, Cur-r. Med. Res. Opin., 3 (1975) 181. 4 R. A. Franklin, A. Aldridge and C. de B. White, Br. J. Clin. Pharmac., 3 (1976) 49’7. Prentice-Hall, Englewood Cliffs, 5 V. G. Levich, Physicochemical Hydrodynamics, N-J., 1962. 6 J. RG%zka and E. H. Hansen, Anal. Chim. Acta, 7E (1975) 145. 7 J. R%%ka and J. W. B. Stewart, Anal. Chim. Acta, 79 (1975) 79. 8 P. Gilgen and P. Rach, Chimia, 32(9) (1978) 345. 9 J. Yamada and H. Matsuda, J. Electroanal. Chem., 44 (19’73) 189. 10 B. Fleet and C. J. Little, J. Chromatogr. Sci., 12 (1974) 747.