Method for anaesthesia by use of a pentapeptide

A method for the induction and/or maintenance of anaesthesia or of neuroleptanalgesia in a mammal comprising the administration to the mammal of an anaesthetic-effective or of a neuroleptanalgesic-effective, non-toxic amount of a peptide of formula EQU H.Tyr.D-Ala.Gly.Phe.D-Leu.NHEt or a pharmacologically and pharmaceutically acceptable acid addition salt thereof.

This invention relates to the medicinal use of a pentapeptide and its acid 
addition salts. 
The present invention more particularly relates to the use of the peptide 
of formula (I) 
EQU h.tyr.D-Ala.Gly.Phe.D-Leu.NHEt (I) 
and its acid addition salts as anaesthetic agents and as neuroleptanalgesic 
agents. 
The abbreviations used herein for amino acids and their radicals are those 
conventional in the art and may be found in, for example, Biochemistry, 
11, 1726 (1972). In the above and throughout the following all references 
are to the L-amino acids and their radicals except in the case of glycine 
and unless otherwise stated. 
The peptide of formula (I) and its acid addition salts, when assessed by a 
number of standard pharmacological procedures, has been found both to 
induce and to maintain anaesthesia in laboratory animals including rats 
and mice. The compounds are effective in this respect when administered by 
a variety of routes including parenteral, for example by intravenous or 
intracerebroventricular injection. Illustrative of the anaesthetic effects 
of the compounds are the following, which should be understood to be 
non-limiting: 
(I) Abolition of the righting reflex. This is characteristic of recognised 
anaesthetic agents such as chloral hydrate 
(2,2,2-trichloro-1,1-ethanediol), urethan (ethyl carbamate) and the 
barbiturates (derivatives of barbituric acid). An animal lacking this 
reflex does not roll over or attempt to regain its normal posture when 
placed on its back. 
(II) Abolition of the pinnal reflex. In this procedure a wire or similar 
probe is introduced into the ear pinna; in the normal (control) animal 
there is a resultant reflux twitch or shake of the affected pinna. 
(III) Abolition of the corneal reflex. In this procedure the cornea is 
lightly touched with a wire or similar; in the normal (control) animal 
there is a resultant reflux blink of the eyelids. This reflex is of 
clinical importance in man in that it is one of the last reflexes to be 
abolished during the induction of general anaesthesia. 
Each of the foregoing effects (i), (ii) and (iii) may be reversed by 
administration of the known narcotic antagonist naloxone 
(1-N-allyl-7,8-dihydro-14-hydroxynormorphinone). However it has been found 
that morphine itself does not abolish the righting reflex in laboratory 
animals such as mice when administered by acute bolus injection in up to 
lethal doses. 
In the acid addition salts of the peptide of formula (I) the activity 
resides in the base and the acid is of less importance although for 
therapeutic purposes it is preferably pharmacologically and 
pharmaceutically acceptable to the recipient. Examples of such suitable 
acids include (a) mineral acids: hydrochloric, hydrobromic, phosphoric, 
metaphosphoric, nitric and sulphuric acids; (b) organic acids: tartaric, 
acetic, citric, malic, lactic, fumaric, benzoic, glycollic, gluconic, 
gulonic, succinic and arylsulphonic, for example p-toluenesulphonic, 
acids. The pharmaceutically and pharmacologically acceptable acid addition 
salts together with those salts which are not so acceptable (for example 
salts of hydrofluoric and perchloric acids) have utility in isolation and 
purification of the bases, and of course the unacceptable salts are also 
valuable in the preparation of the acceptable salts by techniques well 
known in the art. 
The peptide of formula (I) and its pharmacologically and pharmaceutically 
acceptable acid addition salts may be used in the fields of both human and 
veterinary medicine for the induction and/or maintenance of anaesthesia in 
a mammal. 
The peptide or a salt thereof may be administered either alone as the sole 
anaesthetic agent or in combination with one or more other substances 
which may complement and/or supplement its activity. Such additional 
substances may be administered before, simultaneously with or after 
administration of the peptide or salt thereof and in the case of 
simultaneous administration the various agents may be administered either 
as separate doses or as a combination formulation. 
As one possibility the peptide or salt thereof may be administered 
subsequent to administration of a benzodiazepine tranquillizer such as 
chlordiazepoxide (7-chloro-2-methylamino-5-phenyl-3H-1,4-benzodiazepine 
4-oxide), 
diazepam(7-chloro-1,3-dihydro-1-methyl-5-phenyl-2H-1,4-benzodiazepin-2-one 
) and 
oxazepam(7-chloro-1,3-dihydro-3-hydroxy-5-phenyl-2H-1,4-benzodiazepin-2-on 
e). 
As another possibility the peptide or salt thereof may be administered for 
the maintenance of anaesthesia after this has been initially induced by 
the previous administration of another anaesthetic agent, for example a 
barbiturate such as thiopental sodium (sodium 
5-ethyl-5-(1-methylbutyl)-2-thiobarbiturate). 
A particular utility for the peptide of formula (I) and its 
pharmacologically and pharmaceutically acceptable acid addition salts, 
within the field of anaesthesia, is the induction and/or maintenance of 
the state referred to as "neuroleptanalgesia", a condition characterised 
by quiescence, psychic indifference to environmental stimuli, and 
analgesia (see, for example, Dorland's Illustrated Medical Dictionary, 
twenty-fifth edition, published by W. B. Saunders, 1974, at page 1041, and 
"The Pharmacological Basis of Therapeutics", Goodman, L. S. and Gilman, A. 
eds., fifth edition, published by Macmillan Publishing Co. Inc., 1975, 
especially at Chapter 8, pages 97 to 101, all of which is incorporated 
herein by reference hereto). This condition is recognised by clinicians as 
desirable for enabling the performance of procedures such as bronchoscopy, 
X-ray studies, burn dressings and cystoscopy wherein a degree of patient 
cooperation is of value, and a fixed-dose combination comprising the 
narcotic analgesic fentanyl citrate 
(N-(1-phenethyl-4-piperidyl)propionanilide citrate) and the neuroleptic 
agent 
droperidol(1-{1-[3-(p-fluorobenzoyl)propyl]-1,2,3,6-tetrahydro-4-pyridyl}- 
2-benzimidazolinone) has found acceptance for use in such circumstances. 
Heretofore neuroleptanalgesia has been achievable only upon administration 
of such a narcotic analgesic plus neuroleptic drug combination as that 
above mentioned. The peptide of formula (I) and its acceptable acid 
addition salts is thus an important clinical advance and valuable addition 
to the armamentarium of the medical and veterinary professions in alone 
enabling this result, without any additional medication being required. 
In addition to its ability both to induce and to maintain anaesthesia, as 
hereinbefore described, the peptide of formula (I) and its acid addition 
salts has been found to exhibit morphine agonist activity. As generally 
accepted and as the term is used herein, a morphine agonist is a compound 
the biological activity of which mimics that of the natural alkaloid. 
The pharmacological properties and therapeutic uses of morphine are well 
documented in the literature, see for example "The Pharmacological Basis 
of Therapeutics", Goodman, L. S. and Gilman, A.eds., published by The 
MacMillan Company, New York, third edition (1965) especially at Chapter 
15, pages 247 to 266, and "Martindale: The Extra Pharmacopoeia", Blacow, 
N. W. ed., published by The Pharmaceutical Press, London, twenty-sixth 
edition (1972) especially at pages 1100 to 1106, all of which is 
incorporated herein by reference hereto. As is well known however 
(Goodman, L. S. et al., loc. cit, Chapter 16) repeated administration of 
morphine can lead to the recipient developing an addiction to the drug and 
tolerance to its effects and to his manifesting withdrawal symptoms when 
administration is discontinued. For many years therefore research has been 
conducted with the aim of obtaining a compound having the activity 
spectrum of morphine while lacking its disadvantages. 
The morphine agonist properties of the peptide of formula (I) and its 
derivatives as hereinbefore defined include the following, which are given 
solely by way of illustration and should be understood to be non-limiting: 
(A) In vitro 
(i) Inhibition of neurally evoked contractions of the isolated mouse vas 
deferens when tested by the method of Hughes et al (Brain Research, 88 
(1975) 296) (using pulses at 0.1 Hz), the inhibition being abolished by 
the known narcotic antagonist naloxone (1-N-allyl-7,8-dihydro-14-hydroxy 
normorphinone). 
(ii) Reduction of electrically-induced contractions of the isolated 
guinea-pig ileum when prepared for stimulation after the manner of Paton 
(Brit. J. Pharmacol., 12 (1957) 119-127). (Each intestinal segment was 
impaled by the anode and suspended with a 2-3 g load. Stimulus parameters: 
frequency: 0.1 Hz; duration: 0.4 ms; voltage (supramaximal) 30-40 V; the 
contractions were transduced isotonically). 
(B) In vivo 
(i) The compound exhibits analgesic activity, for example it is effective 
in mice in the "hot plate" procedure standard in the art when tested by a 
modification of the method of Eddy, N. B. et al. (J. Pharm. Exp. Therap. 
107, 385 (1953), the compound being administered by 
intracerebroventricular injection, and this activity is abolished by 
naloxone. 
(ii) The compound exhibits antitussive activity, for example when tested in 
guinea-pigs according to the method of Boura et al, Brit. J. Pharmacol, 
39, (1970) 225. 
(iii) The compound exhibits antidiarrhoeal activity, for example it is 
effective in reducing castor oil-induced diarrhoea in rats. 
Because of their morphine agonist activity already alluded to the peptide 
of formula (I) together with its pharmacologically and pharmaceutically 
acceptable acid addition salts may be used in the treatment of mammals in 
the fields of both human and veterinary medicine in any condition where an 
agent with a morphine-like effect is indicated. Specific utilities that 
may be mentioned, by way of example, include the following: 
(1) The relief of pain (analgesia), for example pain arising from spasm of 
smooth muscle as in renal or biliary colic, pain due to terminal illness 
such as cancer, pain in the post-operative period, and obstetrical pain. 
(2) Sedation, for example in pre-anaesthetic medication; tranquillization; 
the induction of sleep, especially where sleeplessness is due to pain or 
cough; and the relief of anxiety in general. 
(3) The suppression of cough. 
(4) The relief of dyspnoea, for example that of acute left ventricular 
failure or pulmonary oedema. 
(5) The induction of constipation, for example after ileostomy or 
colostomy, and the treatment of diarrhoea and dysentery. 
(6) The induction of euphoria and the treatment of depression, for example 
when allied to the relief of pain in terminal illness such as cancer. 
For each of the utilities recited hereinbefore for the peptide of formula 
(I) and its acid addition salts, that is to say, whether for use for the 
induction and/or maintenance of anaesthesia (for example the induction 
and/or maintenance of neuroleptanalgesia) or for use in a condition where 
an agent with a morphine-like effect is indicated (for example the 
utilities specifically indentified hereinbefore under (1), (2), (3), (4), 
(5) or (6)) the amount required of the peptide or acid addition salt 
thereof (hereafter referred to as the active ingredient) will vary with 
the route of administration and with the nature and required extent of the 
desired effect, and will ultimately be at the discretion of the physician 
or veterinarian. In general however for each of these utilities the dosage 
will be in the range 0.0025 .mu.g to 40 mg per kilogram bodyweight of 
mammal, preferably 0.0025 .mu.g to 10.0 mg/kg, more preferably 0.01 .mu.g 
to 4.0 mg/kg and optimally 0.25 to 400 .mu.g/kg (all dosages calculated 
with reference to the peptide base). 
The active ingredient may be administered by any route appropriate to the 
effect to be achieved, suitable routes including oral, rectal, nasal, 
topical (buccal), vaginal and parenteral (including subcutaneous, 
intramuscular and intravenous). It will be appreciated that the preferred 
route will vary with the effect to be achieved and thus for example in the 
relief of obstetrical pain administration directly into the spinal cord 
may be advantageous. 
While it is possible for the active ingredient to be administered as the 
raw chemical it is preferable to present it as a pharmaceutical 
formulation preparation. 
The formulations, both veterinary and for human use, of the present 
invention comprise an active ingredient, as above defined, together with 
one or more acceptable carriers therefor and optionally other therapeutic 
ingredients. The carrier(s) must be "acceptable" in the sense of being 
compatible with the other ingredients of the formulation and not 
deleterious to the recipient thereof. Desirably the formulations should 
not include oxidising agents and other substances with which peptides are 
known to be incompatible. 
The formulations include those suitable for oral, rectal, nasal, topical 
(buccal), vaginal or parenteral (including subcutaneous, intramuscular and 
intravenous) administration, although the most suitable route in any given 
case will depend upon for example the active ingredient and the condition 
to be treated. The formulations may conveniently be presented in unit 
dosage form and may be prepared by any of the methods well known in the 
art of pharmacy. All methods include the step of bringing into association 
the active ingredient with the carrier which constitutes one or more 
accessory ingredients. In general the formulations are prepared by 
uniformly and intimately bringing into association the active ingredient 
with liquid carriers or finely divided solid carriers or both, and then, 
if necessary, shaping the product into the desired formulation. 
Formulations of the present invention suitable for oral administration may 
be presented as discrete units such as capsules, cachets or tablets each 
containing a predetermined amount of the active ingredient; as a powder or 
granules; or as a solution or a suspension in an aqueous liquid or a 
non-aqueous liquid; or as an oil-in-water liquid emulsion or a 
water-in-oil liquid emulsion. The active ingredient may also be presented 
as a bolus, electuary or paste. 
A tablet may be made by compression or moulding, optionally with one or 
more accessory ingredients. Compressed tablets may be prepared by 
compressing in a suitable machine, the active ingredient in a free-flowing 
form such as a powder or granules, optionally mixed with a binder, 
lubricant, inert diluent, lubricating, surface active or dispersing agent. 
Moulded tablets may be made by moulding in a suitable machine, a mixture 
of the powdered compound moistened with an inert liquid diluent. 
Formulations for rectal administration may be presented as a suppository 
with the usual carriers such as cocoa butter, while a suitable formulation 
for nasal administration is nasal drops comprising the active ingredient 
in aqueous or oily solution. 
Formulations suitable for topical administration in the mouth include 
lozenges comprising the active ingredient in a flavoured basis, usually 
sucrose and acacia or tragacanth; and pastilles comprising the active 
ingredient in an inert basis such as gelatin and glycerin, or sucrose and 
acacia. 
Formulations suitable for vaginal administration may be presented as 
pessaries, creams, pastes or spray formulations containing in addition to 
the active ingredient such carriers as are known in the art to be 
appropriate. 
Formulations suitable for parenteral administration conveniently comprise 
sterile aqueous solutions of the active ingredient, which solutions are 
preferably isotonic with the blood of the recipient. Such formulations may 
be conveniently prepared by dissolving solid active ingredient in water to 
produce an aqueous solution, and rendering said solution sterile and 
isotonic with the blood of the recipient. The formulations may be 
presented in unit--or in multi-dose containers, for example sealed 
ampoules or vials. 
Formulations suitable for nasal administration wherein the carrier is a 
solid include a coarse powder having a particle size for example in the 
range 20 to 500 microns which is administered in the manner in which snuff 
is taken, i.e. by rapid inhalation through the nasal passage from a 
container of the powder held close up to the nose. 
It should be understood that in addition to the aforementioned ingredients 
the formulations of this invention may include one or more additional 
ingredients such as diluents, buffers, flavouring agents, binders, surface 
active agents, thickeners, lubricants, preservatives (including 
anti-oxidants) and the like. 
Where the formulation, for human or for veterinary use, is presented in 
unit dosage form, for example those unit dosage forms specifically 
mentioned above, each unit thereof conveniently contains the active 
ingredient (as above defined) in an amount in the range 0.125 g. to 2 g., 
preferably 1.25 g. to 200 mg. and optimally 12.5 g. to 20 mg. (all weights 
calculated with reference to the peptide base). 
The peptide of formula (I) and its acid addition salts may be prepared by 
any of the methods known in the art for the preparation of compounds of 
analogous structure. Thus they may be formed by the sequential coupling of 
appropriate amino acids using either classical methods of peptide 
synthesis or solid phase procedures, or by the initial preparation and 
subsequent coupling of peptide subunits. 
Such reactions may be effected by, for example, activating the carboxylic 
acid group of the ingoing amino acid and protecting the non-reacting amino 
and carboxylic acid groups. Such techniques are standard in the peptide 
art. Details of suitable activating and protecting (masking) groups and of 
suitable reaction conditions (both for the coupling reactions and for the 
removal of protecting groups) giving the minimum of racemisation may be 
found in the following literature, all of which is incorporated herein by 
reference hereto, which is given purely by way of exemplification and 
which is intended to be neither exhaustive nor limiting. 
Schroder and Luebke, "The Peptides" (Academic Press) (1965). 
Bellean and Malek, J. Am. Chem. Soc., 90, 165 (1968). 
Tilak, Tetrahedron Letters, 849 (1970). 
Beyerman, Helv. Chim. Acta., 56, 1729 (1973). 
Stewart and Young, "Solid Phase Peptide Synthesis" (W. H. Freeman and Co.) 
(1969). 
Depending upon the reaction conditions the peptide of formula (I) is 
obtained in the form of the free base or as an acid addition salt thereof. 
The acid addition salts may be converted into the free bases or salts of 
other acids, and the bases may be converted into acid addition salts 
thereof, by techniques well known in the art. 
The peptide of formula (I) and acid addition salts thereof may thus be 
prepared by condensing a reagent (II) 
EQU h--y.sup.1 --oh (ii) 
wherein Y.sup.1 is selected from the radical -Tyr- and a partial radical 
sequence having the radical -Tyr- at its N-terminal end and from thereon 
corresponding to formula (I), with a reagent (III) 
EQU h--y.sup.2 (iii) 
wherein Y.sup.2 corresponds to the balance of the above defined product, 
the reagents (II) and (III) being optionally protected and/or activated 
where and as appropriate; followed if necessary and as appropriate by one 
or both of the steps of deprotection of the product and conversion of the 
product into the base or an acid addition salt thereof. 
It will be appreciated that the peptide of formula (I) may also be prepared 
by reaction of a corresponding peptide alkyl ester, for example the methyl 
ester, with an appropriate monoalkylamine. 
It will be appreciated from the foregoing that what we will claim may 
comprise any novel feature described herein, principally and not 
exclusively, for example: 
(a) A method for the induction and/or maintenance of anaesthesia in a 
mammal, comprising the administration to the mammal of an 
anaesthetic-effective, non-toxic amount of the peptide of formula (I) or a 
pharmacologically and pharmaceutically acceptable acid addition salt 
thereof. 
(b) A method for the induction and/or maintenance of neuroleptanalgesia in 
a mammal, comprising the administration to the mammal of a 
neuroleptanalgesic-effective, non-toxic amount of the peptide of formula 
(I) or a pharmacologically and pharmaceutically acceptable acid addition 
salt thereof.

The following Examples serve to illustrate the present invention but should 
not be construed as in any way providing a limitation thereof. All 
temperatures are in degrees Celsius. 
Experimental Section 
The following abbreviations are used throughout 
HOBT--1-hydroxybenzotriazole 
DCCI--dicyclohexylcarbodiimide 
DCU--dicyclohexylurea 
NMM--N-methylmorpholine 
DMF--dimethylformamide 
Pr--isopropanol 
Pr.sub.2 O--diisopropyl ether 
pe--petroleum ether 
EtOAc--ethyl acetate 
Z--benzyloxycarbonyl 
Bu--tertiary butyl 
BOC--tertiary butyloxycarbonyl 
Bzl--benzyl 
Peptides were examined by tlc on Merck silicagel plates with the following 
solvent systems: 
(1) methylethylketone 
(2) n-butanol:acetic acid:water (3:1:1) 
(3) chloroform:methanol:32% acetic acid (12:9:4) 
(4) chloroform:methanol:0.880 ammonia (12:9:4) 
(5) ethylacetate:n-butanol:acetic acid:water (1:1:1:1) 
(6) chlorofrm:methanol (8:1) 
All amino acids were of the L-configuration unless otherwise stated. 
Optical rotations were determined on a Bendix NPL automatic polarimeter. 
The amino acid compositions of peptide hydrolysates (6N.HCl at 110.degree. 
for 24 hours in evacuated sealed tubes) were determined with a 
Beckman-Spinco Model 120C amino acid analyser or with a Rank Chromostak 
amino acid analyser. 
The following general procedures were used throughout the syntheses of the 
peptides. 
(a) Couplings were carried out in DMF and were mediated by DCCI. 
(b) Amino acid ester hydrochlorides were converted to the free esters by 
addition of the tertiary base, either triethylamine or N-methyl 
morpholine. 
(c) HOBT was added at the coupling stage when fragment condensation 
involved a peptide having an optically active carboxy terminal amino acid 
e.g. coupling with BOC.Tyr.D-Ala.Gly.Phe.OH. 
(d) Couplings were allowed to proceed for 24 hours in the cold room at 
+4.degree. C. 
(e) After coupling, purification was effected by washing with acid and base 
to remove unchanged reactants. 
(f) Alkaline saponifications were carried out in aqueous methanol with an 
autotitrator at pH 11.5 to 12.0 with N.NaOH. 
(g) Benzyloxycarbonyl protecting groups were removed by hydrogenolysis in 
methanol/acetic acid with 10% palladium on charcoal. 
(h) The resulting acetate salts from the above hydrogenolysis were 
converted to the corresponding hydrochlorides by an addition of methanolic 
hydrogen chloride. 
(i) Benzyl protecting groups were removed by hydrogenolysis in methanol 
with 10% palladium on charcoal. 
(j) Tertiary butyl and tertiary butyloxycarbonyl protecting groups were 
removed with N-hydrogen chloride in acetic acid, in the presence of 
anisole to act as a scavenger. Cleavage was allowed to proceed for 60 to 
90 minutes. 
OBu protecting groups on the alcoholic functions of threonine and serine 
were removed with trifluoroacetic acid containing 10% water, cleavage 
being allowed to proceed for 90 minutes. 
EXAMPLE 1 
Preparation of H.Tyr.D-Ala.Gly.Phe.D-Leu.NHEt hydrochloride 
H.Tyr.D-Ala.Gly.Phe.D-Leu.NHEt hydrochloride was prepared according to the 
scheme set out in Table 1. After purification on carboxymethyl cellulose 
the end-product peptide (as the acetate addition salt) was dissolved in 
water. Subsequently one equivalent of 1 M hydrochloric acid was added and 
the resultant hydrochloride obtained by freeze-drying the solution. 
The hydrochloride had the following characterising data by thin layer 
chromatography (Merck silica gel plates and the solvent systems indicated) 
EQU Rf=0.61.sup.2 ; 0.93.sup.4 ; 0.61.sup.5 
and had an optical rotation of 
EQU [.alpha.].sub.D.sup.25 =+46.5.degree. (c=0.5 in methanol) 
Table 1 
______________________________________ 
##STR1## 
##STR2## 
______________________________________ 
*CM52 is the Whatman brand of carboxyl methyl cellulose - peptides are 
eluated in order of basicity by an ammonium acetate concentration 
gradient. 
EXAMPLE 2 
Pharmacological Activity 
H.Tyr.D-Ala.Gly.Phe.D-Leu.NHEt hydrochloride was tested for the following 
activities according to standard pharmacological procedures. 
(A) Analgesia in mice in the hot plate test (modification of the method of 
Eddy, N. B. et al., J. Pharm. Exp. Therap. (1953) 107, 385, the peptide 
being administered by intracerebroventricular injection). 
(B) Antidiarrhoeal activity in the rat. In this procedure rats were straved 
for 24 hours, the peptide then administered either subcutaneously or 
orally followed after 15 minutes by 1 ml castor oil per rat given orally. 
(C) For antitussive testing, guinea-pigs are subjected to an aerosol 
containing 20% citric acid, 30 minutes after administration of compound 
(orally or subcutaneously). The number of coughs during a five minute 
exposure are counted and meaned for six animals per treatment. The method 
is that described by Boura, A. L. A., Green, A. F. and Saunders, I. A. Br. 
J. Pharmac., May 1970, Vol. 39, No. 1, page 225. 
(D) Compounds are tested for anaesthetic effects following 
intracerebroventricular injection in mice. The animals are observed for 
loss of righting reflex (failure to roll over within 10 seconds of being 
turned on their backs), loss of pinnal reflex (failure to flick the ear in 
response to a light touch at the base of the pinna) and loss of corneal 
reflex (failure to blink when the cornea is touched lightly). Loss of all 
three reflexes constitutes anaesthesia. In addition, any other effects are 
noted. If a compound shows a degree of anaesthesia at the starting dose of 
10 .mu.g/mouse, further doses are done and and ED50 calculated. 
Interesting compounds are then tested intravenously in the same manner. 
From the data obtained the respective ED50 figures were calculated (i.e. 
the dose required to elicit the appropriate effect in 50% of the animals). 
N.T.: not tested. 
Table 1 
______________________________________ 
Results expressed as ED50 for H . Tyn . D-Ala . Gly . Phe . D- 
Leu . NHET . hydrochloride 
Anti- 
Anti- tussive 
Mouse hot plate 
diarrhoea mg/kg - Anaesthesia 
.mu.g/mouse-i.c.v. 
mg/kg (rat) 
guinea- Anaesthesia 
.mu.g/mouse 
ANALESIA s.c. pig mg/kg i.v. 
i.c.v. 
______________________________________ 
0.03 10 2.0 20 2.8 
______________________________________ 
EXAMPLE 3 
Pharmaceutical Formulations 
______________________________________ 
(A) Tablet Formulation (20mg/tablet) 
______________________________________ 
Compound of formula (I) 
20 mg 
Lactose 76 mg 
Maize Starch 10 mg 
Gelatin 2 mg 
Magnesium Stearate 
2 mg 
______________________________________ 
Mix together the compound of formula (I), Lactose and Maize Starch. 
Granulate with a solution of the Gelatin dissolved in water. Dry the 
granules, add the Magnesium Stearate and compress to produce tablets, 110 
mg per tablet. 
______________________________________ 
(B) Suppository (5mg/product) 
______________________________________ 
Compound of formula (I) 
250 mg 
Suppository Base (Massa 
Esterinum C) to 100 mg 
______________________________________ 
Melt the suppository base at 40.degree. C. Gradually incorporate the 
compound of formula (I) in fine powder form and mix until homogeneous. 
Pour into suitable moulds, 2 g per mould, and allow to set. 
Massa Esterinum C is a commercially available suppository base consisting 
of a mixture of mono, di, and triglycerides of saturated vegetable fatty 
acids. It is marketed by Henkel International, Dusseldorf. 
______________________________________ 
(C) Pessary (5 mg/product) 
______________________________________ 
Compound of formula (I) 
5 mg 
Lactose 400 mg 
Providone 5 mg 
Meagnesium Stearate 
5 mg 
______________________________________ 
Mix together the compound of formula (I) and Lactose. Granulate with a 
solution of Povidone in 50% aqueous ethanol. Dry the granules add the 
Magnesium Stearate and compress on suitably shaped punches, 415 mg per 
pessary. 
______________________________________ 
(D) Freeze-dried Injection 100 mg/vial 
______________________________________ 
Compound of formula (I) 
100 mg 
Water for Injections to 
2.0 ml 
______________________________________ 
Dissolve the compound of formula (I) in the Water for Injections. Sterilise 
the solution by passage through a membrane filter, 0.2 .mu.m pore size, 
collecting the filtrate in a sterile receiver. Fill into sterile glass 
vials, 2 ml/vial under aseptic conditions and freeze-dry. Close the vials 
with sterile rubber closures secured with an aluminium seal. 
The injection is reconstituted prior to administration by the addition of a 
convenient volumne of Water for Injections or sterile saline solution. 
In the foregoing, the weight of the compound of formula (I) is in each 
instance calculated with reference to the peptide base.