Atropine Absorption after Intramuscular Administration with 2-Pralidoxime Chloride by Two Automatic Injector Devices

KARL E. FRIEDL**’, CHARLES J. HANNAN, JR.*, PAUL w. SCHADLER5, AND WILLIS H. JACOBn*

Received August 26, 1988, from the ’Department of Clinical Investigation and the §Department of Emergency Medicine, Madigan Army Medical Center, Tacoma, WA 98431 -5454, and the nPharmaceutical Systems Project Management Division, US Army Medical Materiel Development Activity, Fort Detrick, MD 21701-5009. Present addresses: *Exercise Ph siology Division, US. Army Research Institute of Environmental bedicine, Natick, MA 01 760, and “Physiology Branch, Academy of Health lciences, Fort Sam Houston, TX 78234.

Acce ted for publication January 26, 1989.

Abstract I2 Compared with manual intramuscular injection, automatic injector delivery substantially enhances drug absorption rate. We exam- ined the effect of two types of automatic injector delivery of two drugs which are components of the standard antidote to anticholinesterase poisoning and which have been previously shown to have a reduced absorption rate when mixed together in a manual injection. In crossover experiments one week apart, 20 nonsmoking healthy young male humans (ages 20-30) were studied after citrated atropine (6.9 pmoli0.7 mL) and pralidoxime chloride (3.5 mmoli2.0 mL; PAMCL) were injected sequentially into a single intramuscular site by either a multichambered autoinjector or a device which delivers the drugs into two separate intramuscular sites (MARK I). Atropine absorption was assessed by the appearance of atropine in the serum and by changes in heart rate, salivary secretion, pupil diameters, and near vision accommodation. Atropine absorption was significantly greater in the first 30 min following injection with the MARK I. The results of this study suggest that: ( 7 ) the MARK I device produces a faster absorption of atropine, probabiy through some combination of its broader dispersal of atropine in the muscle site and its separation of atropine from the PAMCL; (2) salivary secretion may be the most convenient and sensitive marker of atropine action; and (3) the 6.9-pmol (2-mg) dose of atropine delivered by either autoinjector gives near maximal antisialogogue activity in normal male humans. _ _ _ ~ ~ ~~~~

Self-administration of drugs by intramuscular injection has been simplified with the development of automatic injectors (autoinjectors) which drive the needle into the muscle and forcefully deliver their contents following a single action by the user. Such devices have been used in emergency treat- ments to deliver lidocaine, epinephrine, and atropine.1-5 Since the more forceful delivery of an autoinjector gives a broader drug dispersion, this method of intramuscular deliv- ery also increases the rate of drug absorption.6.7 With this increased tissue dispersion, it may be possible to combine several drugs for which absorption is hindered when they are administered by standard manual injection. In the combina- tion of two drugs which comprise the current U.S. Army nerve agent antidote, atropine absorption is delayed by the presence of 2-pralidoxime chloride (PAMCL) at the same injection site,s but a recent animal study suggests that this is not a problem when the two drugs are administered by a multi- chambered autoinjector.9 We report a study in which the absorption of atropine was compared between two injector systems following atropine and PAMCL delivery into the same or separate intramuscular sites.

Experimental Section This study was approved by the Human Use Committee a t Madigan

Army Medical Center and was conducted under a Food & Drug Administration IND (28301) sponsored by the U.S. Army Medical Materiel Development Activity (Fort Detrick, MD).

Study Volunteers-Twenty healthy young (ages 20-30) male

humans were entered to this study after obtaining their informed written consent. All were nonsmokers. The ethnic distribution of subjects was one Asian, one Black, one Filipino, one Hispanic, one Native American, one South Pacific Islander, and 14 Caucasians. The mean body weight was 77.6 * 8.4 (SD) kg (range 65.9-95.5 kg), and the mean percent body fat as determined by a circumference methodlo was 16.3 * 3.4 (range 11.5-23.4). All subjects exercised regularly, averaging 5 d/week of 1-h exercise periods which usually included running.

Autoinjectors-The MARK I device (Survival Technology; Be- thesda, MD), which is currently used by the Army, includes two separate nose-activated injectors which separate from their safety pins when pulled from a single holderbase (Figure 1). The smaller injector (Atropen) delivers -6.9 pmol(2 mg) of atropine in a 0.7-mL volume, with the injection of drug beginning at the moment that the needle emerges from the cartridge. The larger injector (Combopen) delivers -3.5 mmol(600 mg) of PAMCL in a 2.0-mL volume, but drug delivery does not begin until the needle is fully extended. The multichambered device (Combopen M.C.; Survival Technology; re- ferred to in this report as “MCF’”) consists of substantially the same casing and mechanism as the Combopen portion of the MARK I (Figure 1). The same medicaments delivered by the MARK I device are delivered by the MCP device through a single needle. A double plunger system keeps these drugs separated until sequential injec- tion into the muscle site. When activated, the volumes are pushed forward and the atropine is injected first. When the first plunger driving the atropine is completely forward, the PAMCL volume is injected past the first plunger and into the needle through three grooved channels in the plastic casing. Both the MARK I and MCP devices are spring activated and extend 21-gauge needles -2.0 cm into the muscle, delivering drugs in <4 s. The atropine (base) dose delivered by the injectors used in this study was estimated to be 6.6 * 0.2 (SEM) (1.9 mg) and 6.9 * 0.3 pmolidose (2.0 mg) for the Atropen portion of the MARK I and for the MCP, respectively (measured by radioreceptor assay in five injectors from the test batch). The PAMCL delivered by each of the two devices was estimated to be 3.4 ? 0.01 (512 mg) and 3.5 ? 0.04 mmolidose (611 mg) for the MARK I and the MCP; respectively.

Figure 1-The MARK I (below) and the MCP autoinjector devices. The MCP is 14.2 cm long.

728 i Journal of Pharmaceutical Sciences Vol. 78, No. 9, September 1989

0022-3549/89/0900-0728$0 7 .OO/O 0 1989, American Pharmaceutical Association

Data Collectilon-The study was performed in a crossover design with half of the volunteers first injected with the MARK I device and the other half w:ith the MCP device. Volunteers were fasted for 10 h prior to each experiment and received their first meal only after the first 6 h of the experiment. During each experiment the subjects remained in beds which raised the upper body to a 45" angle. One hour before drug administration, an intravenous catheter filled with heparin solution (10 UlmL) was placed in the arm. Three baseline values were obtained in -10-min intervals for heart rate, salivary secretion. pupil diameter, and near vision accommodation. A single baseline blood sample was drawn in this period. The individuals were then injected with one of the two devices. An area of each volunteer's leg was cleaned with alcohol and the injection(s) was then adminis- tered by one of the investigators to the anteriolateral aspect of the upper leg in th8e largest available muscle site. The injectods) was applied perpendicular to the leg and then pushed to activate the needle and initiate the drug delivery. After 10 s it was pulled straight out. In the case of the MARK I, the second injection (PAMCL) was made 1-2 inches away from the first injection site in a caudal-rostra1 plane within 30 s. Sampling was timed from the point of the first injection. In sequence, heart rate data, blood samples, and salivary secretion samples were collected at 3,6, 10, 15,20,30,40, 50,60,90 min and at 2,2.5,3,4,5,6,9 and 12 h. At these times, pupil diameters and near vision accommodation were also measured, except at 15 min and at 3, 6, andl 15 min, respectively.

Physiological Measurements-Heart rates averaged over a 30-s interval were obtained from three-lead ECGs displayed on a heart monitor. Blood samples were obtained after a 2.5-3-mL void volume was drawn andl discarded; catheters were cleared with 2-3 mL of heparin solution (10 UimL) after each blood draw. Plasma samples were stored at -70 "C until assayed. Salivary secretion was measured by stimulation with a drop of lemon juice placed on the tongue for 45 s, after a subject swallowed all the saliva in his mouth; the contents of the mouth were then collected and weighed. Salivary secretion was expressed as a percent of the average of three baseline weights. Mean baseline weight was 1.86 t 0.21 (SEMI g (n = 20), with coefficients of variation 8.4% within individuals (triplicate), 11.0% between experiments (duplicate), and 20.3% between individuals (n = 20). Pupil diameters were obtained by matching calibrated semicircles on a rule to each pupil, taking care not to shade it from ambient light (12-15 foot candles). The amplitude of accommodation was deter- mined by the proximity method11 using a hand-held slide (R. 0. Gulden, Philadelphia, PA). Each eye was tested individually and the measurements were made with subjects wearing full distance cor- rection (their usual spectacles).

Atropine Assay-Atropine was assayed in serum by a previously described modification12 of the radioreceptor method of Metcalfe.13 Atropine concentrations were compared with a standard curve con- structed using atropine sulfate (Sigma, St. Louis, MO) and were expressed in terms of atropine base equivalents [conversion: 1 mol (695 gimol) atropine sulfate = 2 mol(290 glmol) atropine base]. In an atropine range of 1 to 10 nmoliL, the assay had intra- and interassay CVs of 10 and 11.2%, respectively. The limit of detection was -1.0 nmol/L in 10 p.L of plasma.

Creatine Phosphokinase i :CPKtThe CPK was measured in baseline samples and in samples collected at 2, 4, and 6 h after injection using a spectrophotornetric mcthod designed for use with an automated clinical analyzer (DACOS, Coulter Electronics, Hileah, FL).

Statistical Analyses-Salivary secretion, heart rate, pupil size, accommodation, and serum atropine concentration were statistically analyzed as two-factor ANOVAs with repeated measures in a 2 x 18 mixed design (I987 BMDP Statistical Software; Los Angeles, CA). In the ANOVAs ;significant for interactions, specific differences were pursued using paired t test comparisons at each time point, with application of the Bonferroni method for multiple comparisons in the first 90 min. ]Duncan's multiple range test was used to pinpoint changes over time. Times to maximal change were tested by the Mann-Whitney test.

significant differences between injectors between the 6- to 15-min sampling intervals, with a more rapid decline follow- ing injection with the MARK I (paired t test; Bonferroni, p < 0.01). At 10 min, salivary secretion was 24% of baseline following injection with the MARK I, compared with 45% with the MCP (Figure 2). There was no difference in median time to greatest change (MARK I: 40 min; MCP: 50 min) or in the mean minimal levels achieved (MARK I. 8.7 2 1.9 (SEM) %; MCP: 8.3 t 1.6%).

Heart Rate-Heart rate data collected over a 90-min period is shown in Figure 3. Mean heart rates were significantly elevated from baseline a t 10-180-min sampling intervals for MARK I and at 20-180-min sampling intervals for MCP (Duncan's test). Injector differences in heart rate response were significant from the 6-min sampling interval to the 40-min interval, with a greater response after injection with the MARK I (paired t test; Bonferroni, p < 0.01). In these sampling intervals, the mean heart rate difference between injection with the two devices was at least 10 bpm. There was no significant difference in median time to peak (MARK I: 40 min; MCP: 50 min) or in the peak amplitudes achieved (MARK I: 93.6 ? 2.6 bpm; M C P 91.0 _t 2.6 bpm; MARK I- MCP pairwise: 2.6 * 1.7 bpm).

Pupil Size-Pupil diameters were significantly larger than baseline at 30 min (MARK I) and 60 min (MCP) and remained enlarged through the last sampling interval at 12 h (Duncan's test). There were no statistically significant differences in the behavior of pupil diameters when compared by injector. No anisocoria (>1 mm difference) was observed in any subject. Baseline diameters ranged from 3-7 mm among individuals and this wide interindividual variation masked significant differences between injectors. Expressed as change in pupil area, there was a significant difference between responses to injectors only at the 30-min interval, with a greater increase in pupil area following injection by the MARK I (paired t test; Bonferroni method p < 0.05).

Amplitude of Accommodation-Accommodation was sig- nificantly reduced over time after atropine administration (repeated measures ANOVA), although no single sampling interval could be pinpointed as different from mean baseline values. There were no differences between injectors.

Serum Atropine-Mean serum atropine levels were sig- nificantly different between injector devices a t 10 min (Figure 4). In the first sampling interval (3 rnin), seven out of twenty subjects peaked, and another eight peaked at the 10-min

I I v 03610 1520 30 40 50 60 90

Results Time (minutes) Figure 2-Mean salivary secretion expressed as percent of baseline ( ~ S E M ) Over the first period of 90 min after injection with atropine and PAMCL. Inset shows the results over a period of 12 h. Significant paired t test comparisons between the two devices, using the Bonferroni method, are shown as p < 0.05 (*) and p < 0.01 (**).

Salivary Secretion-salivary Secretion was significantly reduced from baseline at the 6-min sampling intervals and did not recover to baseline values until after the 300-min (MARK I) and 360-min (MCP) intervals (Duncan's test). There were

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35 i

30

25

10 c ._

-5

- 1 0 1 ~ 8 I I I 1

0 3 6 1 0 1520 30 40 50 60 90

Time (minutes) Figure 3-Mean change in heart rate (-tSEM) over the first 90-min period after injection with atropine and PAMCL. Inset shows the actual heart rate over a period of 12 h. Significant paired t test comparisons between the two devices, using the Bonferroni method, are shown as p < 0.05 c) and p < 0.01 (").

sampling interval following injection by the MARK I. In contrast, four out of twenty subjects peaked a t 3 min and only 6 had peaked by 10 min after injection with the MCP. The median times to peak were 6 min for MARK I and 25 min for MCP (p < 0.03). The mean of peak levels achieved was not significantly different between injectors (MARK I: 44.6 -t- 4.2 nmolIL; MCP: 38.8 * 3.4 nmoliL). The effect of body weight was tested against peak serum atropine, but this accounted for <3% of the intraindividual variance with a nonsignificmt univariate correlation.

Other Injector Effects-Pain was reported by most sub- jects in this study following injection with either device; however, the pain was not directly attributed to the injection. In most cases, there was no sensation from the injection itself, rather pain began seconds to minutes after the needle was removed. This was most commonly described as an intense cramping in the upper leg. This sensation usually lasted for 2 to 4 h. In other studies, pain has been specifically attributed to the PAMCL component, irrespective of the means of intramuscular administration.14J5

Creatine phosphokinase (CPK) increased linearly over

40

- n I T L -Mark I si C

a, (0

m - 30

n 1

a,

Q .c 20 0 2 c 2 10 a, (I)

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0 3 610 15 20 30 40 50 60 90

Time (minutes) Figure &Mean serum atropine concentrations (t SEM) over the first 90-min period after injection with atropine and PAMCL. Inset shows the results over a period of 12 h . Significant paired t test comparisons between the two devices, using the Bonferroni method, are shown as p < 0.05 (*) and p < 0.01 (**).

time and mean levels increased by -150 UII at 6 h, consistent with previous reports which demonstrate this effect for in- tramuscular PAMCL injection16 but not for intramuscular atropine injection.' There was no difference between injection devices in the CPK response produced.

Three out of twenty subjects injected with both devices developed a welt, -1 to 1.5 inches in diameter, which became noticeable immediately after injection with the Atropen cartridge of the MARK I device only. The welt was slightly discolored (blanched) in two of the three cases, but was not associated with any other symptoms or with any differences in serum atropine levels relative to the remainder of the group. The welts disappeared within -2 h.

Discussion

This study indicates that there are differences between the two injection devices in terms of the rate of appearance of atropine in the serum and the physiological endpoints achieved. These differences are largely confined to the first 40 min following injection. Serum atropine, heart rate, and salivary secretion changed more rapidly following injection with the MARK I device. In order of significant change from baseline measures, atropine levels changed first (3 rnin), followed by salivary secretion (6 rnin), heart rate increase (10-20 rnin), and change in pupil area (30 minj, irrespective of the injection device. The initial decrease in heart rate and the decrease in salivary secretion were both already evident at the first (3-min) interval, confirming that both of these are convenient pharmacodynamic markers of atropine action. Salivary secretion appears to be a more useful endpoint of atropine bioavailability because it is a relatively simple response, unlike the complex temporal mixture of bradycardic and tachycardic effects; it may also be less influenced by other variables such as pigmentation.17

The differences in atropine absorption may be related to the design of the injectors. The Atropen delivers drug as the needle is being moved forward, while the MCP (and the Combopen) does not begin drug delivery until the needle is fully extended. The effect of this difference in drug delivery has been demonstrated in other studies comparing the tissue distribution of radiopaque dye (Figure 5). The Atropen clearly has a broader field of dispersion and this alone would be expected to enhance absorption.7 This advantage may be amplified by the absence of mixing with the higher osmotic concentration of PAMCL, since such mixing has been previ- ously demonstrated to retard atropine absorption.*

The action of the Atropen also probably accounts for the welts seen in three out of twenty subjects injected with the MARK I device. Dermal infiltration produced by early deliv- ery of the drug would be consistent with the blanching effect and the otherwise symptomless and transient nature of this phenomenon, When fired through a 2-mm thickness of typical military clothing layers (a chemical suit and fatigue trou- sers), -3% of the atropine was lost even to the clothing.

It may be possible to match the atropine absorption rate achieved by the MARK I by modification of the MCP device to distribute the two drugs in a different pattern. Unfortu- nately, PAMCL does not store well in contact with metal; therefore, delivery of PAMCL from an Atropen type of device with the needle residing in the solution is currently impractical.]" Atropine absorption may be further maximized with adjustments in atropine volume and concentration, as well as by the addition of spreading drugs to enhance diff~sion.19~20 Other factors may also enhance absorption, including muscular contraction21 and exercise,22 but there appears to be no difference in atropine absorption between injection into the deltoid and the vastus muscles1 even though

730 t Journal of Pharmaceutical Sciences Vol. 78, No. 9, September 1989

Figure 5-Deliveiry of radiopaquc! materiai into a dog leg by a device consisting of the Pdropen (right) arid Combopen cartridges, but arranged to inject simultaneously (unpublished results from contract DAMDl7- 824-2076, U.S. Army Medical Materiel Development Activity).

there are established differences in blood flow23 and the absorption of lidocainez between these two sites.

References and Notes 1. Stuckev. J. G.: Pitt. M.: Slonian. G.Am. J . Cardiol. 1973.32.988. 2. Diederici, K.W.; Wester, IT. A.; Pentz, R.; Siegers, C. P. h e d .

Klin. 1979. 74. 205-208. 3. Sloman, G:,; Hamer, A,; Baker, G.; Hunt, D. Med. J . Australia

4. Koster, R. W.; Dunning, A. J. N . Eng. J . Med. 1985,313, 1105-

5. Lockey, F. D. J . Asthma. Res. 1980, 17, 153-155. 6. Martin, T. EL; Kastor, J . A.; Kershbaum, K. L.; Engelman, K.

7. Sidell, F. R.; Markis, J. E.; Groff, W.; Kaminskis, A. J . Pharma-

8. Sidell, F. R. Clin. Pharm. I’her. 1974, 16, 711-715. 9. Trouiller, G.; Garrigue, H. Etude pharmacologique et pharma-

cocinetique concernant les autoinjecteurs Atropen-Combopen-

1981,4 Apr 1981, 349-351.

1110.

Am. Heart J. 1980, 99, 282-288.

cokinet. Biopharm. 1974,2, 197-210.

Multipen; Centre d’Etudes du Bouchet: France, 1986; NTIS

10. Vogel, J. A.;Kirkpatrick, J. W.;Fitzgerald,P. I.;Hod don, J. A.; Harman, E. A. Derivation of anthropometry based bofy fat equn- tions for the Army’s weight control program, TR No. 17-88; USARIEM: Natick, MA, 1988.

PB86-200110.

11. Stein, H. A.; Slatt, B. J. The Ophthalmic Assistant. Fundamen- tals and Clinical Practice, 4th ed.; CV Mosby: St. Louis, MO, 1983: 1) 145.

12. Prete,-M. R.; Hannan, C. J.; Burkle, F. M. Am. J . Emerg. Med. 1987.5, 101-104.

13. Metcalfe, R. F. Bwchem. Pharmacol. 1981, 30,209-212. 14. Barkman, R.; Edgren, B.; Sundwall, A. J . Pharm. Pharmacol.

1963,15,671477. 15. Haegerstrom-Portnoy, G.; Jones, R.; Adams, A. J.; Jampolsky, A.

Aviat. Space Environ. Med. 1987, 58, 47-53. 16. Sidell, F. R.; Culver, D. L.; Kaminskis, A. JAMA 1974, 229,

1984-1987. 17. Friedl, K. E.; Hannan, C. J.; Patience, T. H.; Mader, T. H.;

Schadler, P. W. J . Auton. Nerv. Syst. 1988,24, 51-56. 18. May, J. R.; Kondritzer, A. A. Effect of the container on the stability

ofaqueous solutions ofpralidoxime chloride; TR No. CRDL-3353; Edgewood Arsenal, MD, 1965; NTIS AD482946.

19. Schriftman, H.; Kondritzer, A. A. Am. J . Physiol. 1957,191,591- 594.

20. Sund, R. B.; Schou, J. Acta Pharmacol. Toxicol. 1964,21, 339- 346.

21. Kety, S. S. A m . Heart J. 1949,38, 321-328. 22. Mundie, T. G.; Pamplin, C. L.; Phillips, Y. Y.; Smallridge, R. C.

23. Evans, E. F.; Proctor, J. D.; Fratkin, M. J.; Velandia, J.; Wasser- Pharmacology 1988,37, 132-136.

man, A. J. Clin. Pharmacol. Ther. 1975,17, 44-47.

Acknowledgments We gratefully acknowledge the contributions of Mr. Troy H.

Patience, Lt. Col. Thomas H. Mader, Ms. Tracey E. Weir, Lt. Col. Robert E. Jones, Mr. Thomas M. Kettler, Dr. Cheng Wan, 1st Lt. Rita Hoop, and Mr. Thomas Pierce to the collection, analysis, and presen- tation of data in this project. We thank the soldiers from the 9th Infantry Division, 2175th Rangers, and Madigan Army Medical Center whose voluntar participation made this study possible. We also thank the technicarreview panel composed of Lt. Col. J u r en von Bredow, Lt. Col. David T. George, Maj. Martin D. Green, Lt. 801. Jill Keeler, Lt. Col. Robert 0. Pick, Col. Brian Schuster, Dr. Frederick R. Sidell, Lt. Col. Gerald Wannarka, and Lt. Col. Timoth B. Weyandt for their helpful suggestions in the review of this stu&. The views, o inions, and findings in this report are those of the authors and sfould not be construed as an official Department of the Army position, policy, or decision. No endorsement of any manufacturer or of any commercial product is implied in this report. This study was supported by a grant from the U.S. Army Medical Materiel Devel- opment Activity, Fort Detrick, MD (Interagenc Order No. 87PP7850). A compilation of data collected in this s t u i is available as a technical report through the National TechnicaTInformation Service (AD-A192260).

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