EDWIN J. FELLOWS. Philadelphia, Pennsylvania. (Received for publication May 12, 1947) 2-Amino-6-Methylheptane Hydrochloride (Code S-Si)

Transcription

1 THE PHARMACOLOGY OF 2-AMINO-6-METHYLHEPTANE EDWIN J. FELLOWS From the Department of Pharmacology, Temple University, School of Medicine, Philadelphia, Pennsylvania (Received for publication May 12, 1947) In 1910 Barger and Dale (1) reported that a number of phenylethyl- and aliphatic-amines exhibited physiological effects which they designated as sympathomimetic. This type of action subsequently was found to be inherent in a number of phenylisopropylamine derivatives (2, 3). While most of the activity in this field has been concerned with studies on aralkylamines, considerable attention has been devoted to aliphatic amines (4, 5, 6, 7, 8, 9, 10, 11, 12) in recent years. Most of the sympathomimetic amines have been appraised primarily for vasoconstrictor, bronchodilator or analeptic activity, however, certain of these agents recently have been reported to exhibit such totally unexpected actions as analgetic and local anesthetic (13, 14, 15, 16). In the course of a systematic study of alkyl- and aralkyl-amines, the following aliphatic derivative was found to exhibit a number of interesting physiological effects: CII, CH-CH, CH,-CH,-CH-CH, / CH, NH,-HC1 2-Amino-6-Methylheptane Hydrochloride (Code S-Si) ANALGESIA. The effect of 2-amino-6-methylheptane (S51)2 hydrochloride on threshold pain stimulus in cats was determined by 1a method similar to that described by Eddy (17). The variations in the normal thresholds by this method were found to be of the order of 25 per cent. In the present studies, significant activity was inferred if thresholds were elevated 100 per cent or more by a given dose. This amount was not considered definitely analgetic unless thresholds were elevated in at least 4 out of 5 cats. As shown in table 1, doses of 3.0, 5.0, 7.5 and 10.0 mgm. per kilogram of body weight3 induced a significant and consistent elevation of threshold pain stimulus without producing excessive side effects. While a marked elevation of threshold pain stimulus was observed after 15.0 mgm., side effects were pronounced. LOCAL ANESTHESIA. The topical local anesthetic activity of S-51 was determined in rabbits using the method previously described (18). As shown in 1This investigation was supported in a large part by the Smith, Kline and French Laboratories Fellowship Fund. 2 In order to facilitate discussion throughout the remainder of the text, it is understood that S-SI refers to the hydrochloride salt of 2-amino-6-methylheptane unless otherwise specified. the remainder of the text it is understood that all doses mentioned are per kilogram of body weight unless otherwise noted. 351

2 352 EDWIN J. FELLOWS table 2, 1.0 per cent solution of S-51 produced anesthesia for an average of 13.1 minutes. A stream of air was passed over a pledget containing the free base of S-51, which is volatile, and subsequently allowed to come in contact with a rabbit s eye. This procedure was found to induce local anesthesia, but did not permit quantitative evaluation. Therefore, the local anesthetic activity of TABLE 1 Analgesia Effect of 2-amino-6-methylheptane hydrochloride (5-51) on threshold pain stimulus, after intraperitoneal injection in cats. mgm. DOSE /bgss. NUMBER/NUMBER ANALG./USED 5/5 10/10 4/5 4/5 5/5 2/5 MAXIMUM INCREASE IN PAIN THR.ESROLD per ceni 378, 388, 194, 294, , 182,63, 140,281,246 63, 218, 93, , 120, 116, , 175, 213, , 121, 145, 119, , 50 Local TABLE 2 anesthesia COMMENTS Topical application of 2-amino-6-methylheptane hydrochloride (S-51), free base, or cocaine hydrochloride to rabbits eyes. COMPOUND NUMBER OF TESTS CONCENTRATION Convulsions-i/5 General. Convulsions-1/10 Salivated-7/10 Panted-5/10 tremors-3/5 No side effects-2/5 Hyperpnea-3/5 Panted-1/5 No side effects-4/5 Dilated Vomited-1/5 Hyperpnea-2/5 Dilated pupils-5/5 pupils-5/5 AVERAGE DURATION OF ANESTHESIA per cent minutes S-5l HCI S-51 (free base) S-51 (free base) Cocaine Hl S-51 (free base) was appraised in aqueous solution and as shown in table 2, in 0.1 and 0.2 per cent concentration produced anesthesia for an average of 7.1 and 14.1 minutes respectively. Under present conditions, 1.0 per cent solution of cocaine hydrochloride exhibited local anesthetic activity for an average of 19.5 minutes.

3 2-AMINO-6-METHYLHEPTANE 353 rue fact that S-51 was found to diminish the excitibility of certain nerve strnettires is interesting because the volatile free base of this substance might be utilized to decrease irritability of certain accessible afferent nerves such as those in the tracheo-bronchiail tree. It appeared desirable, therefore, to determine whether or not the possible usefulness of S-Si would be complicated by marked systemic effects or toxic properties. CiRCULATORY ACTION. The circulatory activity of S-Si was appraised in 14 vagotomized atropinized dogs anesthetized with pentobarbital sodium, and standardized with epinephrine. In these experiments, the pressor activity of millimol of epinephrine was approximated by millimol of S-51,therefore, its intravenous pressor activity is 1/500-1/1000 that of epinephrine. Since tachyphylaxis is readily apparent after S-Si, only one dose of this substance was injected in a given animal in all of the experiments involving determination of epinephrine equivalents. The effect of intravenous injection of 0.5 to 1.0 mgm. of S-Si was studied in 8 vagotomized dogs anesthetized with pentobarbital sodium in which the spleen, a kidney, a section of small intestine and a leg were enclosed in plethysmographs. In these experiments, a negligible decrease in spleen, an increase in kidney, leg and intestinal volume were noted. In 4 experiments on vagotomized dogs anesthetized with pentobarbital sodium, 1.0 mgm. of S-Si caused an increase in rate and amplitude of contraction of the heart as determined by a Cushney Myocardiograph. MISCELLANEOUS EFFEcTS in ANESTHETIZED DOGS. In 15 experiments on dogs anesthetized with pentobarbital sodium, intravenous injection of 0.5 to 1.0 mgm. of S-Si failed to produce a detectable effect on the small intestine, detrusor of the urinary bladder, urine secretion, respiration, va.gus sensitivity, circulatory response to acetyicholine, furfuryltrimethyl ammonium iodide or epinephrine. ISOLATED INTESTINE. The effect of concentrations of 1:10,000 to 1:200,000 of S-Si was determined in 9 experiments on isolated segments of rabbit jejunum. In general, relaxation occurred throughout the entire dose range used. A slight and transient effect was noted in 1: 200,000, whereas marked and prolonged relaxation was observed in 1:10,000 concentration. In two experiments slight augmentation of the intestine was noted during exposure to 1: 50,000 S-5i. MOTOR ACTIVITY. A method of detecting and summating spontaneous activity of rats has been developed and utilized to measure the effect of a number of sympathomimetic amines on the central nervous system (19, 20). Stimulation was inferred if motor activity increased after administration of a given compound. A record of the activity of a normal rat is illustrated in Fig. 1, A. Fig. 1, B is a kymographic tracing which illustrates the effect. of 1.5 mgm. intraperitoneally of amphetamine sulfate on motor activity in rats. The increase in the number of vertical lines in this case indicates for the most part frequent movement about the cage. Doses of 3.0 mgm. of amphetamine sulfate intraperitoneally produced a typical effect (Fig. 1, C). This consisted almost entirely of side-to-side head motion with little, if any, movement about the cage. S-Si was administered in doses of 20, 25 and 35 mgm. (Fig. 1, D, E, and F) intraperitoneally and 150 mgm. (Fig. 1, G) orally. While there is an indication of more activity after

4 354 1DWIN J. FILLOWA.. ISO PI4../ac I FIG.. \Iotor :utivity of Rats. A. Normal activity; 13. Activity after 1.5 mgm. Amphetamine $ulfate intraperit.oneallv; C. Activity after 3.0 mgm. Amphetamine Sulfate intraperitoneally; 1), E. F. Activity after 20, 25 and 35 mgm. of 2-amino-6-meth iheptane hydrochloride (8-51) jut raperitoneall ; G. Activity after 150 mgm. of 8-51 orally. All (loses are rngm. jar kilogram of hudv weight

5 2-AM1NO-6-METHYLHEPTANE 355 S-51 than that observed in normal rats, a typical amphetamine-like effect was never noted. ACU re TOxICITY. Mwe: Doses of 40 to 75 mgm. of S-Si were injected intraperitoneally in mice (table 3). From the observed mortality shown in table 3, the LD60 was estimated to be 59 mgm. TABLE 3 Acute toxicity Intraperitoneal injection of 2-amino-6-methylheptane hydrochloride (S-Si) Mice ANIMAL DOSE mgm./kgm Rats Rabbits Guinea pigs MORTAI.ITY 0/20 1/25 13/20 19/20 0/10 4/10 10/10 10/10 0/6 3/6 4/6 6/6 0/6 2/6 6/6 6/6 6/6 OBSERVATIONS Restlessness-20/20 Depression followed by excitement-25/25 Tremors and convulsions-20/20 Convulsions-20/20 Death 10 minutes after injection-19/20 Tremors-4/10 Increased activity-b/b Convulsions-8/10 Tremors, salivation, increased activity-lo/lo Convulsions and death within 20 minutes Convulsions and death within 10 minutes. Tremors-2/6 Convulsions and tremors-3/6 Convulsions-4/6 Tremors-1/6 Convulsions and death within 7-8 minutes. Slight tremors-6/6 Convulsions-2/6 Excitement-6/6 Convulsions and death 1 hour after injection Convulsions and death 4 hour after injection Tremors, convulsions-6/6 Death minutes after injection. RaLs: S-Si was injected intraperitoneally in doses of 30 to 60 mgm., the mortality ratio shown in table 3 obtained and the estimated LD50 found to be 41.5 mgm. Rabbits: Intraperitoneal injections of 40 to 100 mgm. of S-5i were made in rabbits (table 3). From the mortality data shown in table 3, an estimated LD50 of 44 mgm. was obtained. Guinea Pigs: In these animals 25 to 100 mgm. of S-51 were injected intraperitoneally (table 3). From these data, an LD30 of 39 mgm. was estimated. SUBACUTE TOXICiTY. Each of 10 rats (Table 4) received daily intraperitoneal injections of 20 mgm. (approximately one-half the LD50) of 5-51 for 30 days.

6 356 EDWIN I. FELLOWS At the end of this period these animals showed an average weight gain of 38 grams. Nine rats (Table 4) were given daily oral doses of 100 mgm. of S-51 for 30 days. After each administration all rats evidenced depression which was followed by pioerection and restlessness. These animals showed an average weight gain of 38 grams. Six other rats (Table 4) received oral doses of 75 mgm. of S-5i for 30 days. The only effect noted in these animals was slight depression. The average weight gain for this group of animals was 49 grams. Twenty-five mgm. (slightly in excess of one-half the LD50) of S-Si were injected intraperitoneally in each of six guinea-pigs (Table 4) daily for a 30-day interval. All of the animals in this group evidenced slight tremors during the first few days of the injection period, but after several days had elapsed, slight restlessness was the only symptom noted. Six other guinea-pigs received daily intraperitoneal injections of 20 mgm. (One-half the LD50) of 5-51 for 30 days. TABLE 4 Subacute toxicity Oral and intraperitoneal administration of 2-amino.6-methylheptane hydrochloride (S-Si) daily for 30 days. ANIMAL NUMBER OP ANIMALS ROUTE 01 AD- MINISTRATION DOSE mgm./kgm. WEIGHT AVERAGE kg,,,. BEFORE AVERAGE WEIGHT END OF Rats 9 P.O Rate 6 P.O Rats Guinea pigs Guinea Rabbits pigs Ip. Ip. Ip. Ip kg* All guinea-pigs receiving either the 20 or 25 mgm. dose gained weight throughout the period of administration (Table 4). Each of 8 rabbits received daily doses of 20 mgm. (approximately one half the LD50) of S-51 for 30 days. These animals evidenced little change in weight during the 30-day administration period (Table 4). At the end of the 30 day period of administration all of the animals in Table 4 were sacrificed and sections of heart, lung, liver, kidney, spleen, intestine stomach and the entire brain and cord were removed. Examination of these specimens by Dr. Charles F. Branch revealed that S-51 did not produce tissue damage which could be detected histologically. A number of experiments were carried out in which S-5i (free base) was inhaled by rabbits and rats. The primary purpose of these studies was to determine whether or not this material would produce irritation of the respiratory tract mucosa. Twenty rabbits were immobilized and arranged to inhale fixed doses of 5.0 to 10.0 mgm. of S-51 (free base) daily for a 10-day period. Twelve rats were placed in a gas chamber and subjected to continuous inhalation of

7 2-AMiNO-6-METHYLHEPTANE (free base) plus flavoring agents for 6 hours daily for 30 days. The air flow through the S-51 (free base) mixture was adjusted to a rate of 6 liters per minute in such a manner that 0.4 to 0.55 mgm. of 5-51 (free base) were delivered in each liter of air throughout the entire six hour period. At the expiration of the 10 and 30 day period respectively, the rabbits and rats were sacrificed and the entire trachea, lungs, brain and cord as well as sections of kidney, spleen, intestine and stomach were removed. Examination of these tissues by Dr. Charles F. Branch disclosed that S-51 did not induce histopathologically demonstrable damage. SUMMARY 1. The threshold pain stimulus was elevated in cats after doses of mgm. per kilogram of body weight of 2-amino-6-methylheptane hydrochloride intraperitoneally. 2. Local anesthesia was produced by tropical application of solutions of the hydrochloride or free base of 2-amino-6-methylheptane to rabbit s eyes. 3. Intravenously in dogs anesthetized with pentobarbital sodium, 2-amino-6- methylheptane hydrochloride exhibited i/s0o-i/1000 the pressor activity of epinephrine. 4. As determined with a Cushny Myocardiograph, 2-amino-6-methylheptane hydrochloride was found to cause an increase in cardiac rate and amplitude of contraction. 5. Intravenously in dogs, anesthetized with pentobarbital sodium, doses of 0.5 to 1.0 mgm. per kilogram of body weight of 2-amino-6-methylheptane hydro. chloride had no detectable effect on the small intestine, detrusor of the urinary bladder, urine secretion or respiration. 6. In general, concentrations of 1:10,000 to 1: 200,000 of 2-amino-6-methylheptane hydrochloride caused relaxation of isolated segments of rabbit jejunum. 7. Intraperitoneally in rats, 2-amino-6-methylheptane hydrochloride was found to be almost entirely devoid of a central nervous system stimulating action in non-toxic doses. 8. Acute toxicity data were obtained on 2-amino-6-methylheptane hydrochloride in mice, rats, rabbits and guinea-pigs. 9. Large doses of 2-amino-6-methylheptane hydrochloride were administered daily for 30 days to rats, rabbits, and guinea-pigs. Examination of the tissues removed from these animals disclosed that this compound did not induce histopathologically demonstrable damage. 10. No histological evidence of irritation of the tracheo-bronchial tree mucosa was noted in rabbits or rats which inhaled 2-amino-6-methylheptane free base daily for 10 and 30 days respectively. REFERENCES (1) BAROER, G. AND DALE, H. H., J. Physiol. 41: 19, 1910 (2) CinN, K. K., AND SCHMIDT, C. F., Tins JOURNAL, 24: 339, 1924 (3) ALLEn, G. A., Tins JOURNAL, 47: 339, 1933 (4) TAINTER, M. L., Arch. Internat. Pharmacodyn., 46: 192, 1933

8 358 EDWIN J. FELLOWS (5) ALLE5, G. A., Univ. Calif. Pubi. Pharmacol., 2: 1, 1941 (6) PROETZ, A. W., Ann. Otol, Rhino!. Laryngol., 51: 112, 1942 (7) PROETZ, A. W., Arch. Otalaryngol., 37: 15, 1943 (8) Romw.&x, E., AND SHONLE, H. A., Jour. Am. Chem. Soc., 66: 1516, 1944 (9) JACKSON, D. E., Jour. Lab. Clin. Med., 29: 150, 1944 (10) ROMAN-VEGA, D. A., AND ADRw.n, J., Anesth. & Analgesia, 22: 248, 1944 (11) AHLQUIST, R. P., THIS JOURNAL 81: 235, 1944 (12) AuEs, G. A., Univ. Calif. Pub!. Pharmacol., 2: 15, 1946 (13) KIESSIG, H. J. AND ORZECHOWSK.I, G., Arch. J. Exper. Path., U. Pharmakal., 197: 391, 1941 (14) GOETZEL, F. R., BURRILL, D. J. AND Ivy, A. C., Fed. Proc. 2: 16, 1943 (15) Ivy, A. C., GOETZEL, F. R., H.&iuus, S. C., AND BURRILL, D. J., Quart. Bull. Northwestern Univ. Sch. Med., 18: 298, 1944 (16) KEISSIG, H. J., Scm.un.z, NARKOSE, Anesth. 15: 21, 1942 (17) EDDY, N. B., Tins JOURNAL 33: 43, 1928 (18) FELLOWS, E. J., Proc. Soc. Exper. Biob. & Med. 53: 7, 1943 (19) SCHULTE, J. W., TAINTER, M. L., DILLE, J. M., Proc. Soc. Exper. Biol. & Med., 42: 242, 1939 (20)ScHuLTE, J. W., RE:?, E. C., BACHER, J. A., LAWRENCE, W. S., TAINTER, M. L., Tins JOURNAL 71: 62, 1941.