IE46036B1 - Prostacyclin and derivatives thereof - Google Patents

Abstract

Novel prostaglandin derivatives of the formula Ia are prepared by dehydrohalogenation of a compound of the formula III. The dehydrohalogenation is carried out using alkali metal alkoxide. The symbols in formulae Ia and III have the meaning given in Patent Claim 1. The compounds of the formula Ia and their salts have useful pharmacological properties.

Description

This invention relates to the extraction, isolation and synthesis of a novel prostaglandin derivative, formulations containing it, and its use in medicine.

Prostaglandin endoperoxides (PGG^ and PGH^) are generated from arachidonic 5 acid by a membrane-bound cyclo-oxygenase enzyme system, described by Hamberg and Samuelsson (Biochem, Biophys. Acta. 326, 448 - 461, 1974) and are subsequently transformed to PGfga’ PGE2’ PGD2S or Thromboxane A2· Thromboxane A2 shares with the prostaglandin endoperoxides the important biological properties of contracting strips of rabbit aorta and aggregating blood platelets.

It has now been found that microsomes derived from a variety of mammalian tissues catalyse the enzymic transformation of the prostaglandin endoperoxides to a prostaglandin derivative (hereinafter referred to as prostacyclin) which does not contract strips of rabbit aorta, relaxes strips of rabbit coeliac mesenteric and coronary arteries, has a potent anti-aggregatory action on blood platelets, is a strong vasodilator in whole animals, and has other properties described hereinafter. 46U3G Microsomes derived from rabbit or pig blood vessels such as veins and arteries, and rat stomach fundus produce about 80 - 90% conversion of the prostaglandin endoperoxides. Microsomes derived from rabbit lung tissue and rat pyloric tissue produce 25% conversion of the prostaglandin endoperoxides, whereas those derived from rat kidney, brain, spleen, liver, heart and seminal vesicle tissues produce 5% or less conversion of the prostaglandin endoperoxides.

Prostacyclin is unstable at room temperature in aqueous medium, having a half-life of approximately 10 mins., but its anti-aggregatory activity can be preserved for several days by dissolving the substance in aqueous alkali or in dry acetone and storing at -20°C. On average prostacyclin is 10 - 40 times more potent as an anti-aggregatory agent than PGE-j and 5 -20 times more potent than PGD2, itself a potent inhibitor of platelet aggregation. Prostacyclin also interrupts and reverses the process of platelet aggregation.

Prostacyclin may be prepared biosynthetically by incubating microsomes of fresh mammalian tissue with a prostaglandin endoperoxide (i.e. PGG2 and/or PGH2) and extraction of prostacyclin or a salt thereof from the incubation mixture into an organic solvent from which prostacyclin or a salt thereof is recovered.

Thus, for example, the above process may involve incubation of PGG2 or PGH2 with aortic microsomes in a suitable buffer solution such as Tris buffer, for approximately 2 minutes at a temperature in the region of 22°C. Conversion of the prostaglandin endoperoxides in this case is approximately 85%. - 4 Extraction of prostacyclin may be achieved for example by the addition of cold (0°C) dry diethyl ether to the incubation mixture. The addition of cold ether stops the enzyme reaction and prostacyclin enters the ether phase which may then be separated from the aqueous phase. Evaporation of the ether by standard techniques such as bubbling nitrogen through the solution results in the prostacyclin being left as a residue which may subsequently be resuspended in an aqueous solution for further examination or dissolved in anhydrous acetone and stored at a temperature inthe region of -20°C for future use.

The aortic microsomes employed in the incubation mixture may be extracted from pig or rabbit aortas. Aortas may be frozen solid, preferably by immersing them in liquid nitrogen, and then crushed to form a powder which is resuspended in a suitable buffer solution and subsequently homogenized. The homogenate may then be subiected to sequential centrifugation so as to isolate the microsomal fraction which may be resuspended in deionized water and lyophilized. The incubation mixture vzas shown to have an immediate anti-aggregatory effect by monitoring the aggregation of human blood platelets in a Born aggregometer.

Prostacyclin may also be prepared using other microsomal tissues identified above in substantially a similar manner to that described for aortic microsomes. Prostacyclin formed in the above-identified incubation mixtures has been found to be different from the other metabolic products of prostaglandin endoperoxides so far described.

Its biological properties on isolated tissues, its instability and its potent anti-aggregatory activity shot; that prostacyclin is not PGE2 - 5 or PGF2a· The presence of prostaglandin D2 isomerase in homogenates of several tissues has been described. As prostacyclin is unstable and is a more potent anti-aggregatory agent than PGD2, it cannot be PGD2· Furthermore, PGD2 isomerase is present in the 100,000g. super5 natant, a fraction that did not produce prostacyclin from prostaglandin endoperoxides. Moreover, PGD, isomerase needs glutathione as a cofactor and the incubations were carried out in the absence of cofactors. PGEg, ΡθΡ2αanc* PGG2 were not sut)strates for aortic microsomes and therefore 15-oxo PGs and other products of prostaglandin cata10 holism could not be considered as prostacyclin. Prostacyclin is also unlikely to be a known 15-hydroperoxy PG, firstly because 15-hydroperoxy PGE2 has a contractile activity on rabbit aortic strip and secondly the product(s) of the spontaneous decay of prostacyclin when bio-assayed did not behave like PGE,,, PGF2a> or PGDg. As prostacyclin has an anti15 aggregatory activity it cannot be identical to Thromboxane A2 or B2 as they are pro-aggregatory substances.

Further studies have shown that prostacyclin has the chemical structure shown in formula (I) (R is hydrogen) (see Johnson et al., Prostaglandins 12/6, 915 - 928, 1976).

OH OH (I) Prostacyclin can therefore be named as (5Z) - 5,6 - didehydro - 9 deoxy - 6,9a - epoxyprostaglandin F^.

By the present invention we provide the compounds of formula (1) wherein R is hydrogen or a pharmacologically acceptable'cation (hereinafter referred to as prostacyclin and salts thereof). Salts of prostacyclin include alkali metal salts, alkaline earth metal salts and salts of organic bases. The sodium salt of prostacyclin is especially preferred.

The present invention also extends to solutions of prostacyclin and salts thereof, for example, in an organic solvent such as acetone, as well as solutions, for example, of alkaline pH, which are substantially free from organic material or biological origin.

The above-mentioned sodium salt is preferably isolated in crystalline form and a preferred embodiment of the present invention provides such crystalline material coated with sodium carbonate formed, for example, by washing the sodium salt with aqueous sodium hydroxide followed by air-drying.

The present invention further provides a synthetic process for preparing a compound of formula - 7 4 6 0 3 6 (wherein R is hydrogen or a pharmacologically acceptable cation) which comprises dehydrohalogenating by treatment with a base a compound of formula (III) as 1 2 shown in the reaction scheme below (wherein each of Z and Z is hydrogen or 5 a hydroxy protecting group, e.g. carboxylic acyl or trialkylsilyl such as trimethylsilyl), X is iodine or bromine providing that when X is iodine the base is an alkali metal alkoxide and Y is a group of formula - OR^ (wherein r] is hydrogen, alkyl or a pharmacologically acceptable cation) or a group 2 2 of formula - NHR (wherein R is alkyl) to form a compound of formula (IV) shown below and subsequently (a) when Y represents a group of formula - OR^ 2 2 wherein R is alkyl or a group of formula - NHR wherein R is as defined above) converting the resulting compound into a corresponding compound wherein Y represents a group of formula OR^ in which R^ is hydrogen or a pharmacologically ί o acceptable cation, and (b) (when either or both of the groups Z and Z are 15 hydroxy blocking groups) optionally removing the said blocking group(s).

The base employed in the above dehydrohalogenation reaction is preferably an organic base or an alkali metal alkoxide, e.g. sodium methoxide, the reaction being carried out advantageously under an inert atmosphere.

The compound of formula (III) may be prepared for example from compounds 1 2 of formula (II) shown below (wherein Y,Z and Z are as defined above) by oxidative attack with an appropriate haolgenating agent at the 5,6-double bond with simultaneous or subsequent cyclisation involving the 9-hydroxy group. The oxidative attack may conveniently be carried out by iodine or potassium tri-iodide in the presence of a metal bicarbonate.

The above-described reaction sequence is illustrated in the following reaction scheme:- ? When Z and/or Z in formulae (II), (III) and (IV) are protecting groups, these groups may be removed by methods known in the art, for example base hydrolysis.

When Y in formulae (II), (III)and (IV) above represents the group OR1 in which R1 is alkyl, e.g. methyl, the ester may be hydrolysed 1 2 to a compound of formula (I), the groups Z and Z both preferably representing hydrogen.

Prostacyclin salts may thus be prepared by treating a prostacyclin ester for example the methyl ester, with a strong base such a sodium hydroxide in a suitable solvent and lyophilising the resulting reaction mixture. Prostacyclin itself may be conveniently prepared by base hydrolysis of its corresponding esters or amides in the presence of an appropriate amount of a caustic alkali in an aqueous alcohol or an aqueous tetrahydrofuran medium, and if desired extraction into an organic solvent at low temperature.

Compounds of formula (III) above in which Y represents a group of formula OR1 (in which R^ is as defined above) are described and claimed in our corresponding Application No.19386/76 (Serial No. 1583962). ϋ ϋ 3 6 - 10 Prostacyclin and its salts are useful as intermediates in the synthesis of prostaglandin analogues, and exhibit a potent anti-aggregatory action on blood platelets, and therefore have a particular utility in the treatment and/or prophylaxis of mammals as anti-thrombotic agents.

Prostacyclin and its salts are also useful in the treatment of mammals, including man, to reduce and control excessive gastric secretion, thereby reducing or avoiding gastrointestinal ulcer formation, and accelerating the healing of such ulcers and lesions already present in the gastrointestinal tract.

Prostacyclin and its salts further exhibit vasodilatory action on blood vessels and therefore have a particular utility as anti-hypertensives for the treatment of high blood pressure in mammals, including man.

Platelets can be assimilated into the vascular endothelium or even incorporated into endothelial cells. Biochemical co-operation between platelets and vascular endothelium in the generation of prostacyclin contributes to the repair of vascular endothelium, and prostacyclin and its salts have a further utility in the promotion of wound healing in mammals, including man.

Prostacyclin and its salts may be used whenever it is desired to inhibit platelet aggregation, to reduce the adhesive character of platelets, and to treat or prevent the formation of thrombi in mammals, including man.

For example, they may be used in the treatment and prevention of myocardial infarcts, in the treatment of peripheral vascular disease, to treat and prevent post-operative thrombosis, to promote patency of vascular grafts following surgery, and to treat complications of arteriosclerosis and - η conditions such as atherosclerosis, blood clotting defects due to lipemia, as well as other clinical conditions in which the underlying etiology is associated with lipid imbalance or hyperlipidemia.

Prostacyclin and its salts may also be used as additives to blood, blood products, blood substitutes, and other fluids which are used in artificial extra-corporeal circulation and perfusion of isolated body portions, e.g., limbs and organs, whether attached to the original body, detached and being preserved or prepared for transplant, or attached to a new body. During these circulations and perfusions, aggregated platelets tend to block the blood vessels and portions of the circulation apparatus. This blocking is avoided by the use of prostacyclin or a salt thereof which may be added gradually or in single or multiple portions to the circulating blood, to the blood of the donor animal, to the perfused body portion, attached or detached lb to the recipient, or to two or all of those, for example at a total steady state dose of .001 to 10 mg., per liter of circulating fluid.

It is especially useful to use prostacyclin in laboratory animals, e.g. cats, dogs, rabbits, monkeys and rats, for these purposes in order to develop new methods and techniques for organ and limb transplants.

The amount of prostacyclin or a salt thereof required (hereinafter referred to as the active ingredient) for therapeutic effect will vary with the route of administration. In general a suitable dose for a mammal will lie in the range of 0.01 to 200 mg. per kilogram bodyweight, conveniently 0.01 to ID mg per kilogram. 6 0 3 6 - 12 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. Such formulations are preferably non-aqueous and nonhydroxylic in nature, but alkaline aqueous solutions may be used. Unit doses of a formulation preferably contain between 0.5 mg. and 1.5 g of the active ingredient.

Such formulations, both for veterinary and for human medical use, of the present invention comprise the 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.

The formulations include those suitable for parenteral (including subcutaneous intramuscular and intravenous) administration such formulations, which must of course be sterile, will include a liquid carrier, e.g. Tris buffer and may for example comprise a solution of the active ingredient.

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. 6 Ο 3 G - 13 According to the present invention there are therefore provided:a) the compounds of formula (I) and derivatives thereof protected in the 11 - and/or 15-positions; b) the preparation of prostacyclin comprising: i) incubation of microsomes of fresh mammalian tissue with a prostaglandin endoperoxide, and the extraction of prostacyclin from the incubation mixture into an organic solvent; or ii) its synthesis as hereinbefore described; c) Prostacyclin when obtained by the process described in paragraph (b); d) a pharmaceutical formulation containing Prostacyclin; e) method for the treatment of prophylaxis of thrombosis in a non-human mammal or non-human mammalian tissues comprising the administration of an effective amount of a compound of formula (I). f) method for inducing vasodilation in a non-human mammal comprising 15 the administration of an effective amount of a compound of formula (I); g) method for the prophylaxis or treatment of gastric lesions in a non-human mammal comprising the administration of an effective amount of a compound of formula (I); ΰ Ο 3 © - 14 h) (5Ζ) - 5,6 - didehydro - 9 -deoxy - 6,9α - epoxyprostaglandin F, and its salts la i) method for the promotion of wound healing in a non-human mammal comprising the administration of an effective amount of a compound of formula (I).

The following Examples are provided by way of an illustration of the present invention and should in no way be construed as constituting a limitation thereof.

EXAMPLE 1 Preparation of Prostacyclin Pig aortas were stripped of adventitia, snap frozen in liquid nitrogen, crushed into a fine powder, resuspended in 0.05 M Tris buffer (pH 7.5) (1:4, w:v) and homogenised at high-speed in Polytron (Kimenatic, Lucerne, Switzerland) homogenizer, The homogenate was centrifuged at lOOUxg for 15 minutes and the resulting supernatant centrifuged again at 10,000xg for 5 minutes. The 10,000xg pellet was discarded, while the pellet obtained after centrifugation of the supernatant at 100,000g for 60 minutes was resuspended in deionized water and lyophilized. An average yield of 150 mg. of aortic microsomal powder (51% protein) W per 100 g. of aortic tissue was obtained.

Aortic microsomal powder (5 mg) was incubated with 1 pg of prostaglandin endoperoxide (PGG2 or PGH2) in 0.05 M Tris buffer (pH 7.5) (I ml.) for 2 minutes at 22°C. Enzyme activity was estimated by direct bioassay - 15 of the incubation mixture. After incubation of 2 minutes at 22°C all prostaglandin endoperoxide activity was lost indicated by a lack of contraction of rabbit aortic strips when assayed by the cascade superfusion technique of Vane (Br. J. Pharmac. 23, 359 -373, 1964) indicating 100% conversion of PGGg or PGHg.

The prostacyclin was extracted by the addition of cold dried (over sodium) diethyl ether (I ml) to the incubation mixture which also stopped the enzymic reaction. The prostacyclin entered the ether phase, which was subsequently separated from the aqueous phase.

The ether was evaporated by bubbling nitrogen through it leaving a residue wherein the prostacyclin is present in the free acid or salt form, which was either dissolved in ice-cold 0.05 M Tris buffer (0.5 - 1 ml) and immediately used for platelet aggregation studies or was dissolved in anyhdrous acetone ( 1 ml) and stored at -20°C for future use.

The anti-aggregatory activity of the extracted prostacyclin disappeared on boiling (15 seconds) or on standing at 22°C for 20 minutes.

The anti-aggregatory activity of prostacyclin could be preserved for several days by dissolving the substance in dry acetone and storing at -20°C.

By using rabbit aortas in a similar experiment the same results were obtained. 6 0 3 6 - 16 EXAMPLE 2 Aggregation of platelets in 1 ml of fresh human platelet rich plasma (PRP) was monitored in a Born aggregometer. An immediate anti-aggregatory effect of the fresh reaction mixture of Example! of aortic microsomes with PGH2 or PGG2 was observed. The lowest anti-aggregatory concentration was obtained with samples containing 0.05 - 5 ng prostacyclin/ml.

This activity disappeared after leaving the incubation mixture for 20 minutes at 22°C or boiling for 15 seconds. Aortic microsomes alone (50 pg/ml) could induce aggregation in some PRP. The products of spontaneous degradation of PGG2 (100 ng/ml) had no anti-aggregatory activity.

Diethyl ether extracts of Prostacyclin also inhibited platelet aggregation induced by arachidonic acid and PGGg. The effective anti-aggregatory concentration of prostacyclin after storage in ether was from 1 to 10 ng/ml.

EXAMPLE 3 Prostacyclin was injected intravenously and intra-aortically into normotensive anaethetised rets and the blood pressure and heart rate recorded. Results showed prostacyclin to be a powerful vaso depressor having 5 times the potency of PGE2. - 17EXAMPLE 4 Prostacyclin was found to relax spirally cut strips of coronary artery from the ox. The effect is dose dependent and relaxation can be seen with doses as low as 20 nanograms per 5 ml bath. In isolated hearts from rabbits perfused at constant flow rate by the Langendorf technique, prostacyclin produced coronary vasodilation.

Prostacyclin relaxed strips of coeliac and mesenteric arteries, but was less potent that PGE2EXAMPLE 5 Prostacyclin was shown to inhibit the formation of gastric lesions induced in rats by indomethacin at doses from 62.5 to 250 pg/kg upon subcutaneous injection.

EXAMPLE 6 A stirred solution of PGF2 methyl ester (50 mg) in ether (1 ml) was treated with sodium bicarbonate (115.0 mg; lOmolecular equivalents) and water (1 ml) and then dropwise during 2 hours with aqueous potassium triiodide (0.7 molar; 0.261 ml). After stirring overnight, the reaction mixture was shaken with ether and aqueous sodium thiosulphate; the ethereal phase was separated, washed with water, dried 6 0 3 6 - 18 with magnesium sulphate, and evaporated to leave a yellow gum of b£-iodo-9-deoxy-6C,9a-epoxyprostaglandin methyl ester.

A solution of 55-iodo-9-deoxy^,9a-epoxyprostaglandin F^ methyl ester (100 mg.) in methanolic sodium methoxide prepared from sodium (46 mg.) and dry methanol (0.70 ml.) was set aside under dry nitrogen for 5 hours, then freed from solvent in high vacuum. The residual amorphous solid was washed with benzene, set aside in the air overnight, and stirred with ff.aqueous sodium hydroxide (0.5 ml.) to give a suspension of colourless fine needles. The crystals were collected, washed with a few drops of N aqueous sodium hydroxide, and dried in the air to give the sodium salt of 9-deoxy-6,9a-epoxy-A -prostaglandin F^. The inhibition of arachidonic acid-induced aggregation of human platelets at a concentration of 0.2 ng/ml. by this salt and its instability in water at acid pH, together with further evidence, is compatible with the assignation of the configuration (5Z) - 5,6 -didehydro 9 - deoxy - 6,9a - epoxyprostaglandin F·] sodium salt.

The high-resolution C n.m.r. spectrum of a solution of the crystals in dimethyl sulphoxide-dg showed the expected 20 resonances whose chemical shifts were entirely consistent with the chemical structure established for prostacyclin. No impurity peaks were detected. 6 0 3 6 - 19 EXAMPLE 7 5ξ - Iodo - 9 - deoxy - 6ζ,9α - epoxy prostaglandin methyl ester (500 mg) was stirred with methanolic NaOMe prepared from Na (o.23 g., 10 equivs.) and MeOH (3.5 ml.) under N2 at room temperature overnight; Iff aq. NaOH (2.5 ml.) was added to the yellow reaction solution to bring about hydrolysis of the ester moiety and, after 2 hours, the methanol was evaporated in vacuo at room temperature. The residual aqueous solution gave rise spontaneously to a mass of colourless fine needles of the desired sodium salt (formula (I): r = Na) which was cooled (0°), collected, washed sparingly with Iff aq. NaOH, air-dried, and stored in a stoppered tube; this salt (383 mg.) had \>max (KBR disc) 1692 cm _1 - C = Cj I and twenty C resonances only were observed (at 182.7 (C-l), 158.2 (C-6), 140.0 and 134.3 (C-13,14), 100.7 (C-5), 87.5 (C15), 80.6 and 75.5 (C-9,11), 58.0 (C-12), 49.0, 45.8, 42.4, 41.9, 37.5, 35.8 (C-18), 31.6, 29.9, 29.3, 26.7 (C-19), and 18.4 (C-20) ppm from TMS in DMSO-dg). The product, sodium (5Z)-5,6-didehydro-9-deoxy-6,9a-epoxy-prostaglandin F^ (syn. sodium prostacyclin), thus obtained inhibited arachidonic acid-induced platelet aggregation (human plateletrich plasma) at 1 ng./ml. and its profile of biological activity on the rabbit aorta, rabbit coeliac artery, rat stomach .20 . strip and rat colon conformed with that of sodium prostacyclin obtained by biosynthesis. After air-drying, the salt has a surface coating of sodium carbonate (aa 3.5% by weight) which protects the vinyl ether moiety against carbondioxide catalysed hydrolysis.

Claims (1)

1. WHAT WE CLAIM IS:1. A compound of the formula wherein R is hydrogen or a pharmacologically acceptable cation. 5 2. A compound as claimed in claim 1 wherein R is hydrogen. 3. A compound as claimed in claim 1 wherein R is a pharmacologically acceptable cation. 4. A compound as claimed in claim 3 wherein R is an alkali metcl cation. 5. A compound as claimed in claim 3 wherein R is an alkaline er th 10 metal cation. 4 6 ϋ 3 6 -22 6. A compound as claimed in claim 3 wherein R is an organic base cation. 7. A compound as claimed in claim 3 wherein R is the sodium cation. 8. (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F 1 . 5 9. A solution of (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F-| a substantially free from organic material of biological origin. 10. A solution of (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F.j of alkaline pH substantially free from organic material of biological origin. 10 Π. A solution of (5Z)-5,6-Didehydro-9-deoxy-6,9re-epoxyprostaglandin F 1a in an organic solvent. 12. A solution as claimed in claim 1ί wherein the solvent is acetone. 13. (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F^ sodium salt. 14. Crystalline (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F^ 15 sodium salt. 15. Crystalline (5Z)-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin sodium salt coated with sodium carbonate. 16. (5Z}-5,6-Didehydro-9-deoxy-6,9a-epoxyprostaglandin F-| sodium salt substantially free from an ester of said prostaglandin. 4 6 0 3 6 - 23 17· A process of preparing a compound as defined in claim 1 comprising the dehydrohalogenation by treatment with a base of a compound wherein X is a bromine or iodine, provided that when x is iodine the base is 1 2 5 an alkali metal alkoxide, Z and Z are each selected from hydrogen atoms and hydroxy protecting groups and Y is a group of formula -OR^ (wherein R 1 is hydrogen, C^_ 4 alkyl or a phramacologically acceptable cation) or a group of 2 2 formula-NHR (wherein R is alkyl) and subsequently (a) (when Y represents 11 2 a group of formula - OR wherein R is alkyl or a group of formula - NHR 10 wherein R is as defined above) converting the resulting compound into a corresponding compound wherein Y represents a group of formula OR 1 in which R 1 is hydrogen or a pharmacologically acceptable cation, and (b) (when either 1 2 or both of the groups Z and Z are hydroxy protecting groups) optionally removing the said protecting group(s). 15 18. A process for the preparation of a compound as claimed in claim 1 comprising the incubation of microsomes of fresh mammalian tissue with a prostaglandin endoperoxide, and extraction of a compound, as defined in claim 1, from the incubation mixture into an organic solvent from which prostacyclin or a salt thereof is recovered. - 24 1 2 19. A process as claimed in claim 17 wherein Z and Z are hydrogen. 20. A process as claimed in claim 19 wherein Y is OR^ wherein R^ is alkyl from 1 to 4 carbon atoms, and the resulting ester is hydrolysed to a compound defined in claim 1. 5 21. A process as claimed in claim 20 wherein R^ is methyl. 22. A process as claimed in claim 21 wherein the dehydrohalogenation is effected with an alkali metal alkoxide. 23. A process as claimed in claim 17 wherein the dehydrohalogention is carried out under an inert atmosphere. 10 24. A process as claimed in claim 22 wherein the alkoxide is sodium methoxide, and the resulting ester is hydrolysed with sodium hydroxide to produce a compound of formula (1) wherein R is sodium. 25. A process as claimed in claim 18 for preparing (5Z)-5,6-didehydro9-deoxy-6,9a-epoxyprostaglandinF^ sodium salt comprising the reaction 15 of 55-iodo-9-deoxy^,9a-epoxyprostaglandin F-| methyl ester with sodium methoxide; and reaction of the resulting prostaglandin ester with aqueous sodium hydroxide to yield the desired product in crystalline form. 26. A process as claimed in claim 25 wherein the sodium salt product is washed with aqueous sodium hydroxide and air-dried to provide a coating 20 of sodium carbonate upon the crystals of the sodium prostaglandin salt. - 25 27. A pharmaceutical formulation comprising a compound as defined in claim 1 in association with a pharmaceutically acceptable carrier therefor. 28. A formulation as claimed in claim 27 wherein the compound as 5 defined in claim 1 is (5Z)-5,6-didehydro-9-deoxy-6,9ci-epoxyprostaglandin F l«· 29. A formulation as claimed in claim 27 or claim 28 wherein the carrier is a liquid. 30. A formulation as claimed in claim 29, wherein the carrier is an 10 alkaline aqueous solution. 31. A formulation as claimed in either claim 29 or 30 which is a sterile parenterally acceptable injectable solution. 32. A formulation as claimed in claim 27 or claim 28 wherein the carrier comprisies Tris buffer. 15 33. A formulation as claimed in claim 27 wherein the compound as defined in claim 1 is a pharmacologically acceptable salt of (5Z)5,6-didehydro-9-deoxy-6,9a-epoxyprostaglandin F, . 34. A formulation as claimed in claim 33 wherein the compound as defined in claim 2 is (57.)-5,6-didehydro-9-deoxy-6,9a-epoxyprostaglandin 20 F^ a sodium salt. - 26 35. A method for the treatment of prophylaxis of thrombosis in a non-human mammal or in non-human mammalian tissue comprising the administration to the mammal or the tissue of an effective amount of a compound as defined in claim 1. 5 36. A method for inducing vasodilation in a non-human mammal comprising the adminstration to the mammal of an effective amount of a compound as defined in claim 1. 37. A method for the prophylaxis or treatment of gastric lesions in a non-human mammal comprising the adminstration to the mammal of an 10 effective amount of a compound as defined in claim 1. 38. A method for the promotion of wound healing in a non-human mammal comprising the administration to the mammal of an effective amount of a compound as defined in claim 1. 39. A method as claimed in any of claims 35 to 38 wherein the compound 15 of claim 1 is (5Z)-5,6-didehydro-9-deoxy-6,9a-epoxyprostag1andin F^ . 40. A method as claimed in any of claims 35 to 38 wherein the compound of claim 1 is a pharmaceutically acceptable solution of the anion of (5Z)-5,6-didehydro-9-deoxy-6,9ct-epoxyprostaglandin F^. 41. A method as claimed in any of claims 35 to 38 wherein the compound 20 of claim 1 is (5Z)-5,6-didehydro-9-deoxy-6,9a-epoxyprostaalandin F^ sodium salt. 4 ΰ ΰ 3 6 -27 42. A method as claimed in any of claims 35 to 38 wherein the compound is administered parenterally. 43. A method as claimed in claim 42 wherein the compound is administered intravenously. 5 44. A method as claimed in any of claims 35 to 43 wherein the compound is administered as a solution thereof. 45. A method as claimed in any of claims 35 to 44 wherein the compound is administered in an amount of from 0.01 to 200 mg per kilogram bodyweight of the mammal. 10 46. A prostaglandin as claimed in claim 1 or a pharmaceutical formulation thereof substantially as hereinbefore described. 47. A process of preparing a prostaglandin as claimed in claim 1 substantially as hereinbefore described in Example 1, 48. A process of preparing a prostaglandin as claimed in claim 1 15 substantially as hereinbefore described in either of Examples 6 or 7. 49. A prostaglandin which is an arachidonic acid derivative characterised by the following properties:a) it has an anti-aggregatory action on blood platelets 6. Ο 3 θ - 28b) it does not contract strips of rabbit aorta c) it relaxes strips of rabbit coeliac, mesenteric and coronary arteries. d) it relaxes strips of ox coronary arteries e) it has instability such that its anti-aggregatory action on blood platelets substantially disappears upon standing at 22°C for 20 minutes f) it is a vasodilator in whole animals. 50. A prostaglandin derivative as claimed in claim 49 which is formed by 10 incubating microsomes of pig aortas with PGG 2 and/or PGH 2 endoperoxide(s)