SOME ASPECTS

THE BAEYER-VILLICER REACTION

- by -

JOHN EDVARD BOLLIG-ER. B.So, CSyd.)

A. Thesis

submitted for the degree of

MASTER OF SCIENCE

- at the -

UNIVERSITY OP NEW SOUTH WALES

April, 1963- CONTENTS

Page No. Summary . . .. 1

The Baeyer-Villiger Reaction .. 2 Discussion: .. .. 26

Baeyer-Villiger Oxidation of 2-Bromocholestan-3-one 32 Baeyer-Villiger Oxidation of 2-Bromofriedelin 50

Baeyer-Villiger Oxidation of 2-Chlorocholestan-3-one 53 Baeyer-Villiger Oxidation of 2-Iodocholestan-3-one 54 Baeyer-Villiger Oxidation of Gerin 56

Experimental .. .. 61 Acknowledgments .. .. 87

Bibliography .. .. 88 1

SUMMARY

V/ith a view to synthesising suitable inter­ mediates for intramolecular Darzen's glycidic ester syntheses, certain steroid and triterpenoid a- substituted ketones have been subjected to Baeyer-

Yilliger oxidation. In some cases the oxidation yielded unexpected products and possible mechanisms for their formation are discussed. In other cases the expected products were obtained but readily under­ went an unusual rearrangement. The structures of the rearranged products have been chemically elucidated and the mechanism of the rearrangement is discussed.

This thesis is prefaced by a discussion of the

Baeyer-Villiger reaction. THE BAEYER-VTLLIGER REACTION

The reaction of ketones with peracids to give esters was first observed by Baeyer and Villiger in 1899 . The scope of this reaction, now known as the Baeyer-Yilliger reaction, has since been widely extended and it has found many useful applications in organic chemistry. Among its representative uses, illustrated by many examples in p the literature , are the formation of esters from simple aliphatic or aromatic ketones, formate esters from aldehydes, anhydrides from o—diketones, lactones from alicyclic ketones and enol lactones from a,0-unsaturated alicyclic ketones.

The reaction may be effected under mild conditions and using a wide range of peroxides including neutral, acidic or alkaline hydrogen peroxide, permono- or perdi-sulphuric acid, perbenzoic, monoperphthalic, peracetic or trifluoro- peracetic acids. It is usually carried out simply by dissolving the carbonyl compound in a suitable solvent, adding the peracid and leaving the mixture to stand at room temperature or with mild heating until reaction is complete. - 3 -

With the highly reactive trifluoroperacetic acid the reaction is often complete within a few minutes. The mechanism of the reaction has been the subject of a number of publications 1*3-8,11,12,20^ The meohan- i ism was put forward by Baeyer and Villiger and involved the initial formation of an "oxoxide” intermediate which subsequently rearranged with group migration.

R O7-H 0 Sr" C—0—R'--» R—C — 0—R7 X > R—

Wittig and Pieper^ suggested an electrophilic attack of the carbonyl oxygen by OH* to give an intermediate peroxide which then rearranged.

0—H I > R—C—0—"R +

a 5 Criegee , and Robertson and Waters , postulated nucleophilic attack of the carbonyl carbon by the peracid anion to give a hydroxy perester intermediate. Heterolysis - 4 - of the perester to eliminate an acid anion leaves an electron deficient oxygen atom which is stabilised by the migration of one of the groups.

R .0—H 0-7-K H R—C—0—R * 0 —-0^—X

The validity of this last mechanism was established 6 18 by the elegant experiment of Doering and Dorfman. Ox labelled benzophenone was reacted with perbenzoic acid to yield labelled phenyl benzoate. This was reduced with lithium aluminium hydride to phenol and benzyl alcohol and the distribution of 0X in these products determined. The Baeyer-Villiger mechanism would result in an equal dis- tribution of O1 ft between the phenol and benzyl alcohol l8 whereas that of wittig and Pieper would result in the 0 being exclusively in the phenol. Doering and Dorfmann, how- 1 ft ever, found the 0X to be exclusively in the benzyl alcohol, the result to be expected from the Griegee mechanism. The kinetics of the Baeyer-Villiger reaction have been closely examined by Hawthorne and Emmons'. These workers investigated the trifluoroperacetic acid oxidation of a series of ketones in the presence of trifluoroacetic acid. 5 -

In a non-polar solvent, ethylene chloride, and using a large excess of ketone, the overall rate was found to depend on the identity of the ketone and on the concentrat­ ions of both acid catalyst and peracid but was independent of the ketone concentration. This suggested that the ketone was initially rapidly complexed with one of the other reactants. On mixing ethylene chloride solutions of ketone and trifluoroacetic acid the production of heat and shifts in the ultraviolet spectrum indicated that a complex such as the following was formed

R

In ethylene chloride the rate data were found to fit the second order expression:

-d [CI',CO H) /dt = k2 [CP3C02H] [CP3C02H-R2C0j

In a polar solvent, acetonitrile, complex formation was apparently inhibited due to solvation and the data were found to fit the 3rd. order expression:

-d [CF3C03H]/dt = k3 [CP3C02H] [r2CQ1 [CF3CC>3h] 6

The rates of simple carbonyl addition reactions such as

oximation and semicarbazone formation may be correlated by

a simple linear free energy relationship. For a given

series of ketones a similar relationship should exist be­

tween the free energy of activation for the Baeyer-Villiger

reaction and the free energy of activation for oximation if

the rate determining step of the former reaction were the slow addition of peracid to the carbonyl group. No such relationship was found by Hawthorne. The relative

nucleophilicities of the anions of peracetic and trifluoro- peracetic acids are the reverse of the relative reactivities of the two acids in the Baeyer-Villiger oxidation, the latter being two hundred times faster than the former in the

oxidation of phenyl cyclohexyl ketone . These facts exclude

the peracid addition as the rate determining step and indeed point to its being the acid catalysed decomposition of the intermediate, for the relative reactivities of the tri- fluoroacetate and acetate anions respectively as leaving groups parallel the reactivities of their parent peracids in the Baeyer-Villiger reaction.

The course of the Baeyer-Villiger reaction, i.e. the rate determining elimination of a negative ion to leave a positive centre to which the intramolecular migration of an atom or group then takes place, presents a distinct analogy to a class of ionic 1, 2 shift reactions represented by the - 7 -

Pinacol, Wagner-Meerwein, Hofmann, Lossen, Curtis, Beckmann and other related reactions. In all these reactions the formation of a carhonium ion or atom having a sextet of electrons is followed by group migration, and it has been observed that factors which increase the rates of formation of the positive centres affect the overall rates of these reactions. For example, in the rearrangement of camphene hydrochloride (I) to isobornyl chloride (III), a reaction thought to involve the carbonium ion (II) the reaction rate was found to increase with increasing dielectric constant of the solvent . Compounds such as ferric chloride and mercuric chloride which can coordinate with a chloride ion 22 were found to be excellent catalysts .

The analogy between these reactions and the Baeyer-Villiger reaction is further illustrated by certain other features of it. The migration of asymmetric groups in the Baeyer- - 8 -

Villiger reaction has been investigated and found to proceed with retention of their configurations. Turner studied the perbenzoic acid oxidation of cis and trans 1-acetyl-2-methyl o cyclohexanone0. In both cases the 2-methyl cyclohexyl group migrated with complete retention of configuration to give respectively the cis and trans 2-methyl cyclohexanyl acetates. He obtained similar results with the cis and trans 1-acetyl-2-methylcyclopentanones. Mislow and Brenner found that in the peracetic acid oxidation of optically active methyl a-phenylethyl ketone,

0 II CrH- CH- — Chi > C^H-—CH—0 — C- 6 b I '6 b CH, CH3

the a-phenylethyl group migrated with complete retention of q its stereochemical configuration*^. A further example is the perbenzoic acid oxidation of both the 17a and 17ft isomers of 3a-acetoxy-pregnan-20-one to give respectively the 17a and 17ft isomers of 5ft-androstane-3a,17-diol 10 diacetate . This result is due to the migration with retention of configuration of the two epimeric atoms. In the Pinacol, Hofmann, Beckmann, and other reactions of the beforementioned class of 1, 2 shift reactions, migration - 9 - of asymmetric groups has been observed to occur without 2^-27 Walden inversion or racemisation The relative tendencies of various groups to migrate in the Baeyer-Villiger reaction, generally termed migrat- 11 12 ory aptitudes, have received considerable attention 9

These migratory aptitudes again parallel those observed for various migrating groups in the pinacol-^ and related reactions and may be rationalised by considering the mechanism of the rearrangement in a form that is somewhat simplified but best suited for the present argument. The elimination of an anion from the hydroxy perester inter­ mediate leaves an electron deficient oxygen atom which is stabilised by the migration to it of one of the groups (R, R') with its pair of bonding electrons.

Which of these groups migrates will be determined by the relative electron densities, or nucleophilicities, of the two bonds x and y. If R and R* are alkyl groups the nucleophilicities of these bonds will depend upon induct­ ive effects and will, for example, be greater for bond q 10 -

of the isopropyl group than for bond p of the t-butyl group.

?

Thus one could expect that for alkyl groups migratory aptitudes would increase in the order primary secondary tertiary. Table I (p. 11) contains selected results from the work of various authors11’12’ which illustrate these tendencies. In the case of aryl group migration it may be seen that p-substituents such as the methoxyl group, which tend to render the phenyl group more prone to electrophilic substitution, also tend to enhance its migratory aptitude by increasing the electron density in the migrating bond. 11

TABLE I

R in C6H5-COR Ester Products $£ Alkyl/Phenyl

RCo2-c6h5 CgHj-CO^ Migration ratio

Methyl 90 negl. V.small. -X Ethyl 67 6 7x10 -z n-propyl 65 6 7x10 neopentyl 64 9 1.1x10

benzyl 39 51 1.3

i-propyl 33 63 1.9 cyclohexyl 25 75 3.0

cyclopentyl 44 46 1.1

t-butyl 2 77 39 1-apocamphyl 2.5 97.5 39

p-ch3o-c5h4 X 96

p-ci-c6h4 60 X

p-Br-C6H4 60 X

p no2-c6h4 95 X

x Not isolated.

On the other hand electron withdrawing substitutents,

such as the chloro or nitro groups which decrease the electron availability of the phenyl ring, also decrease its migratory aptitude. 12 -

In the pinacol type rearrangements there is generally a Walden inversion of the group to which migration occurs^ .

If this reaction proceeded via the initial formation of a carbonium ion which then had a finite life before migration, racemisation but not inversion of this centre would occur.

Inversion could only occur if elimination of the anion and migration took place simultaneously. Similarly, in the

Beckmann rearrangement, it is always observed that the group 29 trans with respect to the C=N bond migrates regardless of the relative migratory aptitudes of the hydrocarbon groups of the oxime. The migration and elimination must occur simultaneously, for if the electron deficient nitrogen had a finite existence the amide product would be determined largely by the relative migratory aptitudes of the groups.

Considering the close analogy of the Baeyer-Villiger re­ action to these reactions, and also the predicted instability 13 - of an oxygen atom with a sextet of electrons, it is reason­ able to assume that the elimination and migration processes in the Baeyer-Villiger reaction are also concerted. Also, since the configuration of the migrating group is completely retained in the Baeyer-Villiger reaction it could not at any stage of its migration be completely detached from the intermediate. These considerations now permit a more pre­ cise picture of the Baeyer-Villiger and related reactions to be drawn. The rearrangement may be considered diagrammatically as below.

R R R i y \ ♦ 1 S—C- ---> S —C—0 OH OR J5-H ... X0R,S~ OR

(IV) (VI)

(IV] is the intermediate before migration. As the reaction proceeds the bond b increases in length, R approaches 0, and the energy of the system increases to a maximum. In this state of maximum energy, the activated complex or transition state, conventionally represented by structures 14 - such as V, the oxygen has lost a large share of one of its electron pairs to the departing acid anion, OR'”, and now has a share of the bonding pair of R. The resulting partial positive charge is distributed between the three centres, C, R and 0. It is again possible to rationalise the tendencies in migratory aptitudes by a slightly different argument to the one mentioned before. structural features in R which tend to stabilise or disperse positive charge will thereby lower the energy of the transition state.

When R is an alkyl group this stabilisation is afforded by inductive and hyperconjugative effects which, as pointed out previously, will be greater in tertiary groups than in secondary or primary groups. Hence the energy barrier re­ presented by the transition state will be least for tertiary groups which will consequently migrate more readily than either secondary or primary groups. When R is an aryl group stabilisation of the transition state is afforded by resonance structures such as (VII) . - 15 -

If the para substitutent X is electron withdrawing it will decrease the stabilisation afforded by the phenyl group, whereas if it is electron donating it will increase

this stabilisation and consequently increase the migratory aptitude of the phenyl group. Once the energy maximum represented by the transition state has been reached, the energy of the system goes "down hill” to give the products; an acid anion and the conjugate acid of the ester (VI). It is conceivable that the final step, the loss of a proton from (VI), also occurs simultaneously with the migration to give the ester product from the intermediate (IV) in one concerted step. One notable exception to the reaction course as out­ lined above appears to be the oxidation of some aldehydes to give largely carboxylic acids rather than formate esters, suggesting the preferential migration of a hydrogen atom over a group which has been found to have a high migratory aptitude in another context. For example the peracetic acid oxidations of benzaldehyde and p-methoxybenzaldehyde give quantitive yields of benzoic and p-methoxy benzoic acids respectively^1, In these cases however, elimination of the acid anion from the intermediate is probably accomp­ anied by loss of a proton rather than migration. - 16 -

H

Similar collapse of a bond to oxygen rather than migration of an alhyl group has been observed to occur to a small extent in the oxidation of camphor. (See p.24) An alternative explanation of the migratory aptitudes based on steric rather than electronic factors was proposed 12 by Hawthorne and his co-workers . They suggested that the observed differences in migratory aptitudes were in many cases too small to be rationalised completely on the basis of electronic factors. The argument considers the hydroxy perester intermediate resulting from an unsymmetric- al ketone in which the groups are of significantly different size. There are two possible rotational conformations of this intermediate one of which will be favoured over the other at equilibrium. Thus conformation (IX) which contains the bulky perester group staggered between the hydroxyl and smaller group S will be favoured over (VIII) in which there must be a greater degree of steric interaction. - 17 -

S i •

L

X

(V1IP (IX)

Assuming that the migrating group enters the transition state trans to the leaving acid anion, an assumption which is quite reasonable on the grounds of the foregoing dis­ cussion, it is clear that the sterically favoured conform­ ation IX places the larger group L in a favourable position to fulfil this condition.

This argument has an experimentally documented analogy in the pinacolic deamination of 1,2 diphenyl-l-(p-chloro- phenyl)-2-amino ethanol1^. For this compound there are two racemates pairs) and it was found that in the reaction of the a racemate with nitrous acid the phenyl group migrated while in the reaction with the 0 racemate the p-chloro- phenyl group did. For each racemate there are two rotat­ ional conformations of which one is sterically favoured over the other for the same reasons cited in the previous argument. Thus(X)is the favoured conformation of the 18 -

a racemate and places the phenyl group trans to the leav­ ing diazonium group, ready for migration, while(XI)is the favoured conformation of the |3 racemate and places the p-chloro-phenyl group in the trans position.

(X) (XI)

Both these arguments assume that the rearrangements of the intermediates take place at a rate that is rapid compared with the rate of interconversion of their rotational isomers. In the Baeyer-Villiger reaction there are no such clear cut examples to support this argument and indeed the observation that neopentyl has a migratory aptitude comparable to ethyl and n-propyl groups suggests that it is of little importance.

There is an example of the Baeyer-Villiger reaction however, which despite a number of elaborate attempts to do so, has not been successfully explained by either steric or electronic arguments. It is ironical that this reaction, the oxidation of camphor, was one of the first examples of - 19 -

the Baeyer-Villiger reaction. Baeyer and Villiger1 treated camphor with persulphuric acid in strongly acidic medium and obtained an unstated yield of a-campholide(XII) and no other product.

(XII) (XIII)

From the foregoing arguments this is not the expected isomer as it results from the preferential migration of a primary rather than a tertiary (bridgehead) carbon.

Sauers14 reinvestigated this reaction and found that the peracetic acid oxidation of camphor under strongly acidic conditions gave a 30% yield of a-campholide (XII) together with an oil. The reaction in buffered acetic acid, how­ ever, gave a good (82%) yield of (XIII) and no a-campholide. Treatment of (XIII) with strongly acid conditions led to an oil similar to that obtained in the preparation of (XII) thereby explaining the failure to isolate any (XIII) in the latter reaction. Thus in an acidic medium the overall relative migratory aptitudes of the tertiary bridgehead and 20 primary carbon atoms is about 3*1 whereas apparently no migration of the primary group occurred in buffered med­ ium. Sauers rationalised this in the following manner.

The energy of the transition state for migration as re­ presented byOfl (p. 13) depends, among other factors, upon;

(a) the ability of the migrating group to sustain positive charge; (b) the stability of the newly forming acid anion and (c) the degree of separation of unlike charges. In an acidic medium protonation of the intermediate as in(XIV) is possible and results in its decomposition passing through a transition state, (XV), in which the contribution of (c) to the total energy is eliminated.

s—c—0—0 C‘—R S—C—0. \ I H+ OH '■ OH °\ ‘0—C—R* 6*:l CK..,

(XIV) (XV)

In a buffered medium, however, protonation of the intermed­ iate is inhibited and its decomposition must involve a transition state of higher energy than for the case in acidic medium since this transition state involves the separation of unlike charges. Thus in the acidic medium 21

the relative stabilisations afforded by the migrating groups R and S are of less significance than in the buff­ ered medium and the reaction is consequently less select­

ive in the acidic medium. A similar decrease in select­ ivity with changing conditions has been noted in the

Baeyer-Villiger oxidation of phenyl cyclohexyl ketone. With peracetic acid the phenyl/cyclohexyl migration ratio was lift, while with trifluoroperacetic acid this ratio rose to 22^&. This finding is consistent with the above argu­ ment since the trifluoroacetate anion is more stable than the acetate anion and thus the transition state (V) in which the separating anion, ORf“ is a trifluoroacetate ion will be of lower energy than that in which it is an acetate ion.

This again results in a decrease in selectivity of group migration. 15 The work of Meinwald and Frauenglass indicates that the argument of Sauers is not a complete explanation of the anomaly. They have studied the peracetic acid oxidation of bicyclo (2,2,l)-heptanone-2 (XVI) and bicyclo (2,2,2)- octanone (XVII) in both buffered and acidic media. In all cases they found that bridgehead migration occurs exclusiv­ ely. This and the analagous camphor oxidation was ration­ alised on the basis of the following steric argument. Structures (XVIII) and (XIX) represent the two possible con­ figurations of the hydroxy perester intermediate derived (XVI) (XVII)

0—0 Ac 0-0Ac

(XX)

0-0 Ac

O-OAc

0-OAc

(XX (10 23 -

from camphor. Since the hydroxyl group is less bulky than the perester group (XVIII) should predominate over (XIX) at equilibrium due to steric interaction with the

methyl groups. For the electronically favourable bridgehead migration to occur in (XVIII) the molecule would

have to pass through the sterically unfavourable boat con­

formation (XX). The alternative electronically less favoured migration, however, causes the molecule to pass

through the sterically favourable chair conformation (XXI).

The presence of a strongly acid medium diminishes the

importance of the electronic factor and this steric factor predominates. In the case of (XVI) the equilibrium pos­ ition for the intermediates (XXII) and (XXIII) is reversed, the absence of methyl groups causing the dominant steric factor to be interaction between the perester group and the

Og endo-hydrogen. Bridgehead migration in (XXIII), then, can proceed via the sterically favourable chair conformat­ ion.

This argument assumes that the lactone group is suff­ iciently flexible to be accommodated equally well in either a boat or chair ring. Recent x-ray studieshave shown that this is not so and that the lactone group:

is planar.

Inspection of Drieding stereo models showed that for an 24 - isolated S lactone, or for a £> lactone ring fused cis to either a cyclohexane ring in the boat conformation or to a five membered ring, the condition of planarity for the lactone group imposes a boat conformation on the fe lactone ring. Examples of a S lactone fused cis to a five mem­ bered ring are iridomyrmecin and isoiridomyrmecin in which the boat conformation has been demonstrated by x-ray 1 techniques . For all other systems containing the g lactone, including the two lactones (XIII) and (XIV) derived from camphor, the planar lactone group is accommod­ ated only as the half boat or half chair conformation.

Thus the argument proposed by Meinwald and Frauenglass is no longer tenable.

A further interesting sidelight in the oxidation of camphor in acidic medium is the formation of the dihydroxy lactone (XXV), in about 13# yield1^. This results from an atypical breakdown of the Baeyer-Villiger intermediate involving the collapse of a bond towards electron deficient 25 oxygen rather than group migration. The mechanism is analogous to that proposed for the Beckmann rearrange- 19 ^2 ment of camphor oxime y1J .

/*v O-QAc (XVIII)

.(XXV) DISCUSSION

The original aim of this work was to investigate the usefulness of Baeyer-Villiger oxidation of alicyclic a-halo ketones as a route to suitable intermediates for the previously unattempted intramolecular Darzens glycidic ester synthesis^. The oxidation of an alicyclic a-halo ketone should give predominantly an a-halo lactone due to the diminished migratory aptitude of the halogen substitut­ ed carbon atom. This has previously been discussed at length (p.10), Opening of the lactone to the hydroxy acid, then methylation followed by oxidation of the hydroxyl group to an aldehyde or ketone would give a suitable intermediate which could undergo a Darzen's condensation. For example, using 2a-bromocholestan-3-one (I) as a starting material the following sequence of reactions might be expected to occur. 27 -

(IV)

tPeracid 2.hydrolysis 3.Methytat ion 4.0xrdation 5.Dsrzen's reaction*

Two examples of the Baeyer-Villiger oxidation of an a-halo ketone are recorded in the literature. The first of these is the perbenzoic acid oxidation of 4-bromo-12a- acetoxy pregnan-3,20-dione (V)^. The products of the oxidation, however, were not isolated; instead the crude product was dehydrobrominated by pyridine and a product containing an a,(3-unsaturated lactone group (VII) was isolated in about 20^S yield. This must have been formed from the a—bromo lactone (VI). 28 -

I.Perbenzoie acid 2.Pyridine (V)

Since no other products were isolated and the yield of (VII) was so low, few conclusions can he drawn about this reaction. The other example is the trifluoroperacetic acid oxidation of a-chlorocyclohexanone to give the tv/o isomeric lactones (VIII) and (IX) in 62$ and 6$ yield resp­ ectively*^ .

(VIII) (IX) - 29 -

In the present work 2-halo-3-oxo steroids and triterp- enes were considered to he convenient model compounds by virtue of their easy preparation, the crystallinity of steroid and triterpenoid derivatives and their steric properties. The rigid ring B would constrain the reactive groups of intermediates such as in (III) to lie in close proximity to each other thereby facilitating the Darzens

condensation. The preparation of these halo ketones and

the products resulting from their oxidation by peracids are discussed below.

Bromination of ketones. The brominations of ketones pro­ ceed via their enolic forms and are therefore usually carried out in the presence of acidic or basic catalysts which pro­ mote the enolisation. In complex molecules such as ster­ oids and triterpenes the position and stereochemistry of

the entering bromine atom depend upon an interplay of steric and electronic factors. In the absence of any

steric effects the axial epimer is more stable than the

equatorial one since in the latter there is strong inter­ action between the coplanar C=0 and C-Br dipoles. However

a 1:3 steric interaction between the axial bromine atom and an axial methyl group is sufficient to render the epimer with the equatorial bromine more stable*^. The position of bromination is determined by the direction of enolisat­ ion and this in turn is determined by the relative - 30 -

stabilities of the two possible enols. Cholest-2-ene is

more stable than the A^ isomer*^ and so cholestan-3-one

brominates at position 2. Until recently the view put

forward by Corey-^, that the kinetically determined product of bromination was the axial one, was generally accepted. Thus the bromination of cholestan-3-one was thought to give initially the 2-axial epimer which then epimerised to the thermodynamically more stable equatorial epimer due to 1:3

interaction with the methyl group on C^q. Recent work by Djerassi-^ and his co-workers and by Shoppee^ has indicat­

ed that in the presence of such steric hindrance the kin­

etically determined product is equatorial.

The configuration of the bromine atom may be determined spectroscopically. Thus the infrared absorption of an a-bromo ketone in which the bromine atom is equatorial is

generally 15-17 cm —1 higher than that of both the epimer having an axial bromine and also of the parent ketone.

Jones and his co-workers^ have attributed this frequency

shift to the interaction between the coplanar C-0 and G-Br bonds, the polarity of the latter bond reducing that of the former and thereby suppressing the contribution of CXI) to the resonance hybrid. - 31 -

<----- >

(X)

....m t • t—-----

This effect is small for the axial arrangement where the tv/o bonds are more or less perpendicular to each other. In the ultraviolet spectrum on the other hand, the situation is reversed, an axial bromine atom causing a bathochromic shift of 25 mp while the absorption maximum of the equator­ ial epimer is at about the same wavelength as the parent ketone.

Baeyer-Villiger Oxidation of 2a—Bromo-5a-oholestan-3-one( I). A range of peracids could have been used to effect the Baeyer- Villiger oxidation of the bromocholestanone and other halo ketones however trifluoroperacetic acid appeared to be the most suitable. This peracid has been found to oxidise cyclohexanone and cyclopentantone instantaneously and in almost quantitative yields to the corresponding lactones^ and was also successful in the oxidation of a-chlorocyclo- hexanone^ (p.25). The conditions under which it is used are important. The reagent can be prepared by simply mix­ ing hydrogen peroxide with trifluoroacetic acid, the - 32 -

equilibrium^:

H202 + CF3C02H;=^ CI^CC^H + H20

being rapidly established. This reagent is suitable for some oxidations but was found to be unsuitable for the oxidation of ketones as the free hydrogen peroxide reacts

to give chiefly the ketone peroxide^2. The reagent is best prepared in situ by mixing 80-90# hydrogen peroxide with excess trifluoroacetic anhydride. Transesterificat­ ion of the ester products with trifluoroacetic acid has been observed to produce some 5-10# alkyl trifluoroacet- ate^2’12. This has been overcome by the addition of di­ basic sodium phosphate which removes most of the acid as soon as it is formed^. The oxidation of 2a-bromocholestan-3-one in chloroform solution containing disodium hydrogen phosphate by tri- fluoroperacetic acid is a mildly exothermic reaction. Attempts to recrystallise the product from methanol at first produced gels. However, after making several attempts, progressively adding more solvent, a beautifully crystalline compound (XII) was eventually isolated in 75# yield. The Beilstein copper wire test showed that this compound did not contain bromine and its analysis was consistent with it having either of the formulae C27H4602.CH30H or C28H4803. Treatment of (XII) with hot - 33 -

alcoholic alkali gave a monocarboxylic acid (XIII) the analysis of which was consistent with either of the form­

ulae ^2qH^qO^ or C^H^O^.CH^OH. Methylation of (XIII) with diazomethane gave back the original compound (XII) thereby establishing it to be a methyl ester. This was confirmed by a positive hydroxamic acid test and a meth- oxyl analysis. It seemed most likely then that the

empirical formula of the methyl ester was C28H48°3 and that of the acid was

The loss of bromine under acidic conditions and the gain of a methoxyl group could not be rationalised by any

possible reaction with trifluoroperacetic acid. These facts, however, suggested that a product was initially form­ ed and that this then reacted with methanol in the recrystall- isation step. This was confirmed by comparing the crude product before and after treatment with methanol by thin

layer chromatography on silica gel, and by comparative infrared spectroscopy in the carbonyl region. The oxidat­

ion reaction, then, was repeated and the crude product care­

fully chromatographed on silica gel. A bromine containing

compound (XTV) C^yH^^O^Br, was isolated. On treatment with methanol (XIV) gave the methyl ester (XII) and on treatment with base it gave the carboxylic acid (XIII).

Structure of (XII) Since compound (XII) had the molecular formula ^28^48°3 an<* Possesse<* a methyl ester group, the - 34 - functionality of one oxygen atom had still to he deter­ mined. An answer to this problem was at first suggested by infrared spectroscopy. The spectrum of (XII) in carbon tetrachloride showed two well resolved, strong bands of equal intensity at 1757 and 1727 cm”1 respective­ ly while that of (XIII) in halocarbon oil mull showed two —1 such strong bands at 1734 and 1719 cm respectively and a weaker band at 1760 cm”1. This suggested that the third oxygen was a carbonyl function. A rationalisation of the formation of such structural units could be made from the results of the analogous Baeyer-Villiger oxidation of a-chlorocyclohexanone. This gave a small (6?&) yield of the lactone (IX) resulting from the migration of the chlor­ ine substituted a carbon atom. Treatment of this lactone with base gave adipic semialdehyde (XV) and with ethanolic

2,4 dinitro phenyl hydrazine solution, the 2,4 dinitro phenyl hydrazone of the ethyl ester of adipic semialdehyde.

a x) CKV) - 35 -

By analogy, it seemed feasable that compound (XIV) had

structure (XVI) and that this was rearranged by base to (XVII).

The ranges in which the carbonyl stretching frequencies of

the pertinent groups generally occur are", aldehydes, 1740-

1720 cm ; normal saturated esters, 1750-1735 cm”'; car­

boxylic acids, 1725-1705 cm_i( dimer) , 1760 cm*"1 (monomer)6^.

The band of (XII) at 1757 cm"-1 is high, however all the

other bands can be accomodated by structures (XVIIa) and

(XVIIb) respectively.

Chemical evidence, however, did not confirm the presence

of an aldehyde nor of a second carbonyl. Neither (XII) nor

(XIII) yielded a 2,4-dinitrophenylhydrazone derivative and

attempted oxidation of (XIII) under strong conditions gave

the starting material back unchanged. Lithium aluminium

hydride reduction of (XII) gave an alcohol (XVIII)

which showed no infrared absorption in the carbonyl region.

Acetylation of the alcohol with either acetic anhydride/ pyridine or refluxing acetic anhydride gave a monoacetate - 36 -

(XIX) ^29H50°3’ showed no infrared absorption in the hydroxyl region. Chromic acid oxidation of the alcohol

(XVIII) gave a carboxylic acid identical with {Till).

The foregoing experiments are summarised below in the flow sheet.

(I) C2?H45OBr

(xiv) G27nA3o^T

a

(XIII) c27h46o3 « 13 ....* - (XII) c28h48o3

\K

(XVIII) C27H48°2

7.

4*

(XIX) c29h50o3

1. Trifluoroperacetic acid 2. Base 3* Methanol

4. Diazome thane 5* Lithium aluminium hydride

6. Chromic acid 7. Acetic anhydride - 37 -

The most consistent explanation of these results is that the third oxygen atom of compounds (XII) and (XIII) is an ether.

Structure of (XT/) Since the structure (XVI) seemed no longer tenable for compound (XIV) the most obvious alternat­ ive structure (II) was considered. A proof of this was seen to lie in its reduction by chromous chloride to give the known lactone (XX) which has been prepared by the 45-7 Baeyer-Villiger oxidation of cholestan-3-one^ '.

Chromous chloride has been used in the reduction of a-bromo and a,0-epoxy ketones to the ketone and 0-hydroxy or 48 a,0-unsaturated ketone respectively . It has also been found successful in the reduction of an a,0-epoxy lactone 49 to the a,0-unsaturated lactone , however no example of its use in the reduction of an a-halo ester or lactone is re­ corded. The reductions are thought to proceed in the 49 following manner. ' 38 -

fiBt H"1’ H

-C^-C- C —C G — C i Hi 1 II H 0^ H 0 H 0

— G—C^C — 0— f -C— 0=0 — 0— —» — c~c—c—o- /I ^ 15 II CK OH 0' 0 H+

The initial product is an enol which subsequently ketonises and since the enolic forms of esters and lactones are less stable than those of ketones, one could expect that the reduction of an a-bromo lactone would proceed less readily than that of an a-bromo ketone. Indeed it was found that in acetone, the solvent generally used for these reductions, the reduction of (XIV) did not proceed satisfactorily while in acetic acid it apparently went to completion to give a fair yield of a compound which did not contain bromine.

Such reactions involving the separation of charged species are generally facilitated by polar solvents which can lower the energy of the charged species by solvating them.

The oxidation of cholestan-3-one was originally 45 carried out by Ellis and Gardner using ammonium per­ sulphate. They separated by fractional crystallisation a major product mp 201-2° and a minor product mp 184-6°. - 39 -

The reaction was later repeated by Burkhardt and Reich- stein^ using perbenzoic acid. They isolated a lactone mp 187°, [a] D + 1.2° as the major product and establish­ ed its structure to be (XX) by converting it to dihydro Diel's acid. Recently Hara, Matsumoto and Takeuchi"^ reported that the oxidation of cholestan-3-one gives a difficulty separable mixture of the two isomeric lactones

(XX) and (XXa). However, these authors give no experi­ mental data or physical constants in their publication. In the present work cholestan-3-one was oxidised both with tri- fluoroperacetic acid and by the method of Burkhardt and Reichstein and in both cases the major product was a lactone nip 199-200°, [a + 1.2°. Thin layer chromatography on silica gel did not indicate that this material was a mix­ ture . It was shown to be identical with the compound obtained from the chromous chloride reduction of (XIV) by comparison of their nujol mull infrared spectra, melting points and by mixed melting point. Hence compound (XIV) may be assigned structure (II).

Barbier-wieland degradation of (XII). The formation of a compound such as (XII), containing methyl ester and ether groups, by the reaction of (XIV) with methanol seemed difficult to rationalise reliably at this stage. It was therefore decided to attempt to elucidate the structure of (XII), the most obvious method being a degradative one, - 40 - namely the Barbier-Wieland degradation. In this degrad­ ation the addition of a Grignard reagent, phenyl magnesium "bromide, to an ester gives a "bisphenyl carbinol. De­ hydration of this and ozonolysis of the resulting olefin gives an aldehyde or carboxylic acid if the carbon a to the methyl ester group is primary and a ketone if it is secondary. In the present case, the formation of a ketone or lactone would most probably mean that the ester group were attached directly to a ring. The reaction of (XII) with phenyl magnesium bromide etherate was carried out in anisole at 100°. It gave an almost quantitative yield of a compound (XXI), Q 2,9^56® 29 corresponding to the addition of two phenyl groups and the loss of CH^O. This compound showed hydroxyl absorpt- —1 ion in the infrared at 3566 cm and no carbonyl absorption It was dehydrated by refluxing for six hours in acetic acid containing 16% acetic anhydride. Longer reaction times led to the formation of side products, as shown by thin layer chromatography. The dehydration product (XXII),

^39H54^ wkich was chained in 70% yield, showed double bond _ i absorption in the infrared at 1618 cm . The case of the dehydration step is due largely to the high degree of stabilisation afforded the intermediate carbonium ion by electron release from the phenyl groups. 41 -

& further factor is the resonance stabilisation of the

conjugated product; hence the choice of a Grignard re­

agent such as phenyl magnesium bromide. The troublesome step in degradations of this type is often the ozonolysis reaction. Ozone adds to most ethylenic double bonds to give a primary product which can then rearrange to the required "isozonide” structure or to 50 a wide variety of side products . The extent of the formation of these side products depends largely on the structure of the ethylenic compound but is thought to be minimised by conducting the ozonisation in a polar solvent and at a low temperature. The decomposition of the isozonide is generally considered to be best achieved by catalytic hydrogenation when aldehyde or ketonic products are desired51. The olefin (XXII) was therefore ozonised 42 - in dilute ethyl acetate solution at about -40° and the resulting ozonide decomposed by catalytic hydrogenation over Adam's platinum oxide catalyst at room temperature. The product of the ozonolysis was a neutral compound (XXIII) which dissolved in aqueous alcoholic alkali and was reprecipitated on acidification. Its

infrared absorption spectrum had a peak at 1735 cm and it did not yield a 2,4-dinitrophenyl hydrazone derivative and showed no absorption in the ultraviolet. These facts together with the empirical formula indicate that (XXIII) contains a six membered ring lactone group. Hence the methyl ester (XII) from which it was derived must have

contained a six membered ring cyclic ether in place of the steroid ring A, with the methyl estdr group attached to one of the carbon atoms adjacent to the ether oxygen. It then

follows that the partial structures of the olefin (XXII)

and of the alcohol (XXI) are as follows. - 43 -

The ultraviolet spectrum of (.XXII) tended to confirm this structure. The spectrum shows absorption maxima at 212 mia, (e = 1.73 x 10^) and 265 mil, (e = 1.54 x 10^).

Systems containing the phenyl group in conjugation with another chromophore give rise to absorption in the ultraviolet in which three main components can be dis­ tinguished; a high intensity band of benzenoid absorption between 200-215 mu; a second high intensity band due to the new conjugated system and usually in the region 230- 260 mil and low intensity benzenoid absorption in the region

270-280 m|i52,53 1,1-Diphenylethylene absorbs at 250 m^, (e = ll,00Cp^ and the spectra of several steroid substituted 44 -

1,1-diphenylethylenes are recorded^55# These all show intense absorption maxima at 250 mu with one interesting exception in which coplanarity of the diphenyl ethylene system, a necessary condition for resonance, is sterically hindered . This results in a lowering of both the wave­ length and intensity of the absorption maximum. The some­ what higher wavelength of the absorption maximum of (XXII) is attributable to the auxochromic effect of an ether oxygen in conjugation with the diphenylethylene system and is thus further evidence for its structure. The following 5 p analogous example supports this view^ ;

CH-.-CH = CH-C0„H X v 204 mu (6=104} J) iliclX •

CH -0-C = CH-C02H Xmax. 234 mu (e=1.4xl04)

ch3

The auxochromic effect is apparently due to interaction of the unshared electron pair on oxygen with the electrons of the diphenylethylene system, thereby extending the con­ jugation.

Attempted Synthesis of (XXIII). As a conclusive proof of its structure an attempt was made to synthesise the lactone (XXIII). Cholestanol was oxidised with permanganate to 2,3-seco-5a-cholestane-2,3-dioic acid (XXIV)' , the reaction presumably proceeding through the enol of cholestan-3-one. - 45 -

Only a small (20$) yield of the dicarboxylic acid was produced together with considerable amounts of intract- ible gummy acidic material. Variation of the reaction temperature failed to increase the yield. Distillation of the dicarboxylic acid with acetic anhydride gave A-nor-cholestan-2-one (XXV)^.

-> o~c(

lA • (XXIV) (XXV) I

The Baeyer-Villiger oxidation of A-nor-cholestanone should yield either one or both of the two isomeric lactones;

(XXVI) (XXVII) - 46 -

In practice only one lactone, G26iiA-402* was isolated and thin layer chromatography and fractional crystallisation

indicated that it was homogenous. This lactone was shown by mixed melting point and comparative nujol mull infrared

spectroscopy to he not identical with (XXIII) and could

therefore he isomeric with (XXIII). However, since thin

layer chromatography did not separate it from (XXIII) and in view of the previously noted work of Hara, Matsumoto and Takeuchi^ in v/hich the Baeyer-Villiger oxidation of 5P and 5a-3-oxo-steroids was found to give difficulty separable mixtures of isomeric lactones*the possibility remains that it is a mixture of (XXVI) and (XXVII).

Although the structure of (XXIII) was not completely resolved, the partial structures of (XII) and (XIII) and

their derivatives were now known and it was thus possible

to rationalise the formation of (XII) and (XIII) from (XIV) and thereby to infer their structures. It is possible to envisage two alternative routes whereby (XIV) could re­

arrange under the influence of methanol and base to give (XII) and (XIII) respectively. The first of these assumes that the combined effects of the carbonyl, the ether oxygen,

and C-Br dipoles produces an appreciable partial positive

charge on rendering this centre prone to nucleophilic attack. Such an attack would result in a breaking of the

lactone ether bond at to give a negatively charged - 47 - oxygen atom which would then eject the bromine on by a rearside attack.

Although this mechanism is plausible for attack by hydroxide ion it is less likely in the case of the weakly nucleophilic methanol. A more likely explanation is that the rearrangement is initiated by the tendency of the highly polarised C-Br bond to ionize. The rearrangement would then involve the elimination of bromide ion and the migration to C£ of the lactone ether bond at in one concerted step.

(XIV) (XII) - 48 -

It has been previously noted that the preferred conformation of the lactone group is planar (p.23) and an inspection of a Drieding stereomodel of (XTV) showed that the equatorial C-Br bond is in the plane of the lactone group. Presum­ ably the planarity of the system would facilitate the simultaneous ionisation of the bromine atom and migration of the ether oxygen. The mechanism is envisaged as being concerted for if a carbonium ion were formed then one might expect an a,P-unsaturated lactone to be produced.

The alternative concerted elimination of hydrogen bromide is presumably not as favourable as the migration of oxygen because the system: H

is not planar. Br It is clear from the above considerations that lactone

(XXIII) from the Barbier-Wieland degradation of (XII) has structure (XXVII). It is then reasonable to assign structure (XXVI) to the lactone formed from the Baeyer-

Villiger oxidation of A-nor-cholestanone.

The infrared spectrum of (XII). Since the structure of the methyl ester (XII) follows from that of (XXIII) it is now possible to reconsider the misleading infrared spectrum of (XII). Josien and Callas^ have studied the infrared spectra of a series of a-halo esters and that of one 49 -

a-phenoxy ester in carbon tetrachloride solution. In the

case of the a-monohalo and a a,a-dihalo esters and also of

the a-phenoxy ester they observed a splitting of the

carbonyl frequency while for the a,a,a-trihalo ester only

one carbonyl band, corresponding to the higher frequency

bands of the mono- and dihalo-derivatives, was observed.

They attributed the carbonyl frequency splitting to rotational isomerism. For example, two rotational con­

formations of allyl a-phenoxy acetate;

o6h5-o-gh2 C02-CH2-CH CH.

— 1 —1 which shows two bands at 1743 cm and 1769 cm respect­

ively, can be represented as below i.e. looking along the

o>, C6H,rQ- •H

A

The higher frequency band can be attributed to the conform­ ation A in which there is the greater degree of interaction between the C-0 and C=0 dipoles^ ^cf. p. 30). Similar- - 50 - ly, in the case of (XII), two such rotational conformat­ ions can he envisaged and are represented helow, i.e. looking along the bond which attaches the methyl ester group to ring A.

This explanation of the carbonyl frequency splitting of

(XII) in carbon tetrachloride solution is further support­ ed by the solid state (nujol mull) spectrum which shows only one band at 1730 cm. .

Baeyer-Villiger oxidation of 2-Bromofriedelin. Since equatorial bromoketones appeared to be unsuitable start­ ing materials for the preparation of the required inter­ mediates for the Darzen's condensation, the axial analogues were considered. Inspection of Brieding stereo­ models of seven membered ring lactones showed that the axial bond a to the lactone group was well out of the plane of that group. Hence an axial a-bromo lactone derived 51 - from an axial a-bromo ketone may not undergo the facile rearrangement undergone by ^XIV) . Compounds such as

5-bromocholestan-6-one and 7-bromocholestan-6-one are known to have stable axial bromine atoms^ but it was thought to be desirable to have a model more closely analogous to 2-bromocholestan-3-one. Both axial

2- and 4-bromo-5a-cholestan-3-one have been prepared^ but are readily epimerised to the equatorial epimers.

The most suitable model, then appeared to be 2-bromo- friedelin in which the bromine has a stable axial orient­ ation due to the absence of a methyl group on C^q, a unique feature, since all other natural triterpenes and steroids have a C^q methyl group.

Friedelin was extracted from industrial granulated 66 cork and brominated in chloroform solution in the pres­ ence of hydrogen bromide to give the 2-a-(axial) bromo derivative-^. Confirmation of the axial orientation of the bromine atom comes from the infrared absorption at 1710 cm , identical with that of the parent ketone, and from the ultraviolet absorption at 310 mu which corresponds to a bathochromic shift of 25 mb as compared with the parent 60 ketone . Attempts to oxidise this bromo ketone using trifluoroperacetic acid under the usual conditions,i.e. in chloroform solution containing disodium hydrogen phosph­ ate, gave back the starting material unchanged. The use of stronger conditions, such as long periods of refluxing - 52 - or leaving the reaction to stand for several weeks in the dark and at room temperature, also failed to give an appreciable amount of product. An inspection of Gatalin stereomodels of bromofriedelin and the hydroxy perester intermediate derived from it suggested that the slowness of the reaction might be due to steric hindrance. When the reaction was conducted in the absence of disodium hydrogen phosphate, thus allowing strong acid catalysis by trifluoroacetic acid, a fairly rapid reaction occurred but gave an intractable oily product. The infrared spectrum of this material showed strongly hydrogen bonded hydroxyl _ \ absorption and carbonyl bands at about 1720 and 1790 cm indicating that it was probably largely the trifluoro- acetate^ (XXIX). This could have resulted from trans­ esterification of the required bromo lactone (, XXVIII) by trifluoroacetic acid - 53 -

Baeyer-Villiger oxidation of 2a-chlorocholestan-3-one.

It now seemed possible that a suitable approach to the desired intermediate might lie in the oxidation of chloro ketones. For example the oxidation of 2ct-chloro-5a- cholestan-3-one should give the chloro analog of (XIV).

Since the tendency of the C-Cl bond to ionize is much less that of the C-Br bond, it may be possible to treat the chloro analogue (XIV) with base and obtain the required a-halo hydroxy acid without rearrangement to (XIII). The a-chloro lactone resulting from the oxidation of a-chloro cyclohexanone was opened with carbonate to give such a hydroxy acid*' , however, this example is not strictly analogous to the chloro anolog of (XIV) since in the for­ mer the chlorine atom is axial and the steric constraint which facilitates the rearrangement of (XIV) is absent.

The chlorination of cholestanone has been shown to be best achieved with t-butyl hypochlorite^2. Since this reagent is also suitable for the oxidation of secondary alcohols to ketones, the chlorination and oxidation were combined to give 2a-chlorocholestan-3-one from cholestanol in one step. The mechanism of the chlorination probably involves an electrophilic attack of the enol double bond by chlorine cation in an analogous manner to bromination^.

The trifluoroperacetic acid oxidation of 2a-chloro- cholestan-3-one proceeded readily to give a product (XXX), - 54 -

C27H45O2GI, which was assigned the following structure.

Cl

(XXX)

The similarity of the specific rotations and infrared carbonyl absorptions of (XIV) and (XXX) tended to confirm this structure. Compound (XXX) was quite stable to treat ment with methanol, however, on treatment with an aqueous alcoholic solution of potassium carbonate it gave an acid identical with (XIII).

Baeyer-Yilliger oxidation of 2a-Iodocholestan-3-one. The third member of the series of 2a-halo-5a-cholestanones

2a-iodocholestan-3-one, has been shown to behave somewhat differently to its bromo- and chloro- analogs. On treat­ ment with collidine the bromo and chloro ketones dehydro- halogenate to give cholest-l-en-3-one while iodocholestan- one is reduced to cholestanone, the collidine apparently acting in this case as an electron source . lodo- cholestanone also shows unusual ultraviolet absorption - 55

^max ^58 (log e = 2.9)^. It was therefore of

interest to complete the series by subjecting iodocholest- anone to Baeyer-Villiger oxidation. It was prepared by

treating a solution of bromocholestanone in acetone with sodium iodide , the bromine apparently undergoing

nucleophilic replacement by iodide ion. Upon adding the

trifluoroperacetic acid reagent to a chloroform solution

of iodocholestanone an intense violet colour, character­

istic of solutions of iodine in chloroform, was immediately produced and cholest-l-en-3-one (XXXII) was isolated in

about 50$ yield. A possible explanation of this seemed to be that the iodine undergoes nucleophilic displacement by the trifluoroperacetate ion and the resulting trifluoro- peracetate (XXXI) then eliminates trifluoroperacetic acid to give the stable conjugated product.

(XXX!) (XXXII) - 56 -

If this were so then the more strongly nucleophilic peracetate anion should also effect the reaction. This was not found to he the case. A further possibility is that the peracid dissociates to give 0H+ ions which then react as follows.

H ck

A series of brief test tube reactions tended to discount both these explanations as the violet colour so readily produced by trifluoroperacetic acid was not produced by peracetic acid nor by 87^ hydrogen peroxide containing p-toluene sulphonic acid to promote its dissociation i.e.

H202 + H+;=±H20 + 0H+

.As it did not appear to be closely related to the present line of investigation, this interesting result was not investigated further.

Baeyer-Villiger oxidation of Cerin. A further approach to the required intermediate was the Baeyer-Villiger oxidation of cerin (2^hydroxyfriedelin (XXXIII)) to the 57 -

hydroxy-lactone (XXXIV, R=H). Replacement of the equa­ torial 2-hydroxyl group by halogen would give the 2-halo lactone (XXXV) which could then he converted by the sequence of reactions outlined on page 26 to a suitable

KU«w

(XXXI I! 5

isolated together with friedelin by the chloroform extract­ ion of cork° . It was subjected to Baeyer-Villiger oxid­ ation by peracetic acid both in refluxing chloroform solut­ ion and in sodium acetate buffered acetic acid solution at room temperature. Surprisingly, in both cases 2,3-seco-

friedelan-2,3-dioic acid (XXXVIII) was obtained in 70$ yield together with a 20$ yield of a compound C^qH^qO^.

This compound reacted with one equivalent of alkali and was reprecipitated on acidification. It showed infrared

absorption at 3417 cm and 1706 cm and was thus assigned structure (XXXIV, R=H). The dicarboxylic acid was identified by mixed melting point and by comparison of its nujol mull infrared spectrum with that of an authentic - 58 -

sample prepared by the chromic acid oxidation of cerin6^. One explanation of this result is that the hydroxyl substituted G^ migrated in preference to to give

(XXXVI). This was converted by acid to the aldehydo acid (XXXVII) and the aldehyde group then oxidised to give friedelin dicarboxylic acid (XXXVIII).

This course is unlikely as the Baeyer-Villiger oxidation of friedelin gives mainly the lactone resulting from the

preferential migration of the secondary atom^^. The

hydroxyl group on would be expected to diminish the migratory aptitude of that centre. A more probable explanation is that the hydroxyl group is oxidised to a

ketone more rapidly than the normal Baeyer-Villiger oxidat­

ion of cerin to give (XXXIV, R=H). The resulting fried-

elan-2, 3-dione then undergoes a normal Baeyer-Villiger

oxidation to give the acid anhydride (XLI) which is sub­ sequently hydrolised. The formation of the anhydride probably proceeds by addition of the peracid to one of the - 59 - carbonyl groups to give an intermediate (XXXIX) which then rearranges with migration of the other carbonyl carbon. The migratory aptitude of the carbon atom carrying the carbonyl oxygen is enhanced since the trans­ ition state for the migration of that centre can be partially stabilised by a polarization of the ft electrons of the carbonyl bond. This transition state is represent­ ed diagramatically by (XL).

(XXXIX) (XL) (XL!)

Acetylation of the cerin prior to the Baeyer-Villiger reaction would prevent its oxidationto the dione and the

Baeyer-Villiger oxidation would then give mainly the acetyl lactone (XXXIV, R=Ac) if the foregoing argument were correct. Cerin was acetylated with acetic anhydride/ pyridine 70 and subjected to trifluoroperacetic acid oxid­ ation. Recrystallisation of the product from methanol gave a compound ^32^52^4 wkich on treatment with alcoholic alkali followed by acidification gave a compound identical 60 with lXXXIV, R=H) . Acetylation of (XXXIV, R=H) with refluxing acetic anhydride gave a compound identical with the Baeyer-Villiger oxidation product of cerin acetate. Thus the structure of the latter compound must be (XXXIV, R=Ac). Alkali treatment of the un- crystallisable mother liquors gave a further amount of

(XXXIV, R=H) making the total yield of this material obtainable from the Baeyer-Villiger oxidation of acetyl cerin about 70$. This tends to confirm the postulated scheme for the formation of the 2,3-seco acid (XXXVIII) from cerin. EXPERIMENTAL 61 -

GENERAL

Melting Points. Melting points were determined on a

Kofler apparatus (C. Reichert, Vienna) and are uncorrected.

Specific Rotations. Specific rotations were determined on

a Lippich polarimeter (Schmidt and Haensch) at room temp­

erature and, unless otherwise specified, in chloroform

solution and in a 1 decimetre tube. The light source was a sodium vapour lamp.

Infrared Spectra. Infrared measurements were determined with a Perkin Elmer421 grating spectrophotometer and have a minimum accuracy of t 2 cm”1. Ultraviolet Spectra. Ultraviolet absorption spectra were determined in ethanol solution in a 1 cm cell on a Perkin Elmer model 137 automatically recording spectrophotometer.

Column Chromatography. The glass columns used were chosen

to he of such a size that the height of the packing was about

ten times the diameter of the column and the total height

of the column twice the height of the packing. After packing the columns were set aside for at least twelve hours before use. Silica gel columns were prepared in the following manner.

The required amount of B.D.H. "Silica gel for chromatog­ raphic absorption" was placed in a separating funnel, covered with n-hexane and shaken until completely wetted - 62 -

with the hexane* The column was filled with hexane, the stem of the separating funnel placed under the liquid

surface in the column and the silica gel slurry allowed to run in while the column was tapped lightly with a piece of

wood. A litre of hexane was then passed through and the column set aside.

Alumina columns were prepared by pouring the dry alumina powder (Peter Spence, gradeH) at a slow even rate into the

column which was filled beforehand with the solvent to be used as the first eluent. The column was tapped lightly with a piece of wood during this operation and a litre of

solvent then passed through it.

Thin Layer Chromatography. Five glass plates, each 10x20 cm, were laid end to end against a straight edged piece of wood fixed to the bench. Merk Silica gel G (7g) and water (14 ml) were made into a thin even paste which was poured onto the first plate and then spread in an even film onto all the plates. The spreader used was simply a rectangular brass block (10x2x1 cm) raised 0.3 mm from the surface of the glass plate by shoulders at its ends. After spreading, the plates were air dried for 5 minutes and then placed in an oven at 120° for 30 minutes. The material to be chromatographed was dissolved in a little chloroform and spotted onto the plate about 1.5 om from its end by means of a glass capillary. The chromatograms were - 63 - developed in closed glass jars containing just sufficient solvent to cover the lower 5-7 mm of the plate. The solvent front was allowed to travel 10-15 cm and the plate then removed, allowed to dry and sprayed with a solution of chlorosulphonic acid in acetic acid Is3. After spray­ ing the plates were heated in an oven at 120° for 10-20 minutes to colour up the spots. Adam’s Platinum oxide catalyst. platinum residues (3g) were dissolved in aqua regia and the solution evaporated to dryness. The residue was redissolved in hydrochloric acid and again evaporated to dryness. The residue was dissolved in water (10 ml) filtered, and a saturated solution of ammonium chloride added until precipitation was complete. The precipitated ammonium chloroplatinate was mixed with A.R. sodium nitrate (30g) and fused for 30 minutes. On cooling the solid mass was dissolved in water, the platinum oxide (2.4g) filtered off, washed briefly with water and dried in an oven at 100°. 5a-Gholestan-3(3-ol. A solution of Cholesterol (B.D.H. laboratory reagent grade) recrystallised twice from 95$ ethanol (50g), and perchloric acid (10 drops, 70$) in ethyl acetate (1100 ml) was hydrogenated at 40-50° and atmospheric pressure over Adam's catalyst (0.8g). The reaction was complete in 50 minutes. The solution was treated with potassium hydroxide solution (1 ml; 50$) - 64 -

filtered, evaporated to about half its volume and set aside for 2 days. The product crystallised in large laths (33g) mp. 140-141°. The mother liquors were evaporated to dryness and recrystallised from 95# ethanol to yield a further 7.5g material mp.139-140°. The total yield of cholestanol mp.139-141° was 8l#. 5a-Oholestan-3-one. A solution of cholestanol (30g) mp.

140-141° in benzene (300 ml) was added slowly and with cooling to a solution of sodium dichromate (41g), acetic acid (30 ml) and concentrated sulphuric acid (54 ml) in water (180 ml). The mixture was agitated vigorously at room temperature for 6 hours. The benzene solution was separated and washed with water (2x60 ml), 5# potassium hydroxide (1x120 ml) and water (2x60 ml) and then dried over anhydrous sodium sulphate. The benzene was evaporat­ ed under reduced pressure and the residue recrystallised from 95# ethanol to yield cholestan-3-one (26g, 87#) as colourless prisms mp.128-129°. 2-a-bromo-5a-cholestan-3-one(1). A solution of cholestan- one (15g) in glacial acetic acid (600 ml) at room temperat­ ure was treated with ten drops of 5# hydrogen bromide in acetic acid. This was followed by the dropwise addition of a solution of bromine in acetic acid (1.05 M; 36 ml).

A flocculent white precipitate formed almost immediately. The mixture was set aside overnight, the precipitate

(13.4g) collected, washed with a little acetic acid follow- - 65 -

ed byioe cold ethanol and recrystallised from chloroform: ethanol 1:4. 2a-bromo-cholestan-3-one separated as need­ les (10.9g, 60$) mp.169-170.5°. Trifluoroperacetic acid reagent.

(cp3co)2o + h2o2—> gp3co3h + cf3co2h

2.10g trifluoroacetic anhydride (0.01 moles) + 0.312g 87$ x/v hydrogen peroxide (0.008 moles) ---> 0.008 moles

trifluoroperacetic acid. Method of preparation: Trifluoroacetic anhydride (3 ml)

was added dropwise during 10 minutes to a cooled, stirred

suspension of 87$ hydrogen peroxide (0.3 ml) in chloroform

or methylene chloride (6-10 ml). The resulting solution

contained approximately 0.008 moles of trifluoroperacetic acid and was used immediately.

2 —carbomethoxy-3-oxa-5a-cholestane (XII). A solution of trifluoroperacetic acid (0.064 moles approx.) in chloro­ form (50 ml) was added dropwise during 20 minutes to a vigorously stirred solution of 2a-bromo-cholestan-3-one (20g, 0.043 moles) in chloroform (260 ml) containing sus­ pended anhydrous disodium hydrogen phosphate (15g). To­ wards the end of this addition the temperature rose to 40°G. Stirring was continued for a further 80 minutes and the temperature maintained at 30-40°. The inorganic material was extracted into water and the chloroform layer successive­ ly washed with dilute sodium bisulphite solution, water and 66 dried over anhydrous magnesium sulphate. The chloroform was evaporated under reduced pressure to leave a light orange glass which was dissolved in refluxing methanol (200 ml) and set aside overnight. The product separated as needles (I4g, 75$) mp.1120. Further recrystallisation from methanol gave pure (XI3), as needles mp.H4°j jof] -5.5°;

0max 1757 cm-1, 1727 cm_1(CCl4), 1730 cm_1(nujol).

Pound: 0,77.83; H.11.32; 0,12.0; 0CH,, 6.62$

0,77.79; H,11.30*

G28H48°3 recluires 0,77.72; H,11.18; 0,11.09 ;0CH^ , 7.5$

Hydroxamic acid test: (XII) (10 mg), pyridine (0.5 ml)and hydroxylamine hydrochloride (10 mg) were heated together at 100° for 2 hours. Dilution with water gave a colourless precipitate which was redissolved in ethanol (2 ml), acid­ ified and treated with 5$ ferric chloride solution. A strong violet colour was produced. 2-carboxy-3-oxa-5a-oholestane (XIII) . (XII) (500 mg) was refluxed for 2 hours in N/2 alcoholic potassium hydroxide solution. Acidification with hydrochloric acid followed by dilution with water gave a colourless precipitate

(490 mg) which was recrystallised repeatedly from methanol to give plates mp.224-5°; jo] p -10.8°; 'O (halocarbon oil mull) 3514; 3442 cm-1 (OH) 1760, 1734, 1719 cm-1 (0=0). Pound: C, 74.92; H.11.10; Equivalent weight 418,414.

03 .CH30H (monocarboxylic acid) required 0,74*66; - 67 -

H, 11.10$;equivalent weight, 450. Hethylation of (XIII). Nitrosomethylurea (1.7g) was added in small portions to a cooled mixture of ether

(25 ml) and potassium hydroxide solution (45$ w/v, 3.5 ml). The ether layer, which contained approximately 0.4g dia­ zomethane, was decanted and dried over potassium hydroxide pellets. It was added to a solution of (XIII) (280 mg) in ether containing 15$ methanol (80 ml) and the mixture set aside overnight at room temperature. The solvent was

evaporated and the residue redissolved in ether (50 ml),

extracted with 3N sodium carbonate, water, dried over anhydrous magnesium sulphate and again evaporated to dry­

ness. The residue was recrystallised from methanol to

yield colourless needles (250 mg) mp.113-114°; [a] ^ -5°, which showed no depression in melting point on admixture with (XII). The nujol mull infrared spectrum was identic­ al with that of (XII).

2a-bromo-4-hydroxy-3>4-seco-5a-cholestan-3-oio acid lactone (XIV). The procedure for the preparation of (XII) was re­

peated on a 2g quantity of 2a-bromo-cholestan-3-one. The

crude product (1.8g) however, was not dissolved in methan­

ol but in carbon tetrachloride (5 ml) and adsorbed onto a

column of silica gel (lOOg) made up in hexane. Elution with hexane:benzene 4:1 (4.51 gave unreacted starting material (0.190g) while elution with hexane:benzene 1:1

(5.51) gave (XIV) (lg). This was recrystallised from 68 - acetone to give small needles mp. 192-193°; [a] ^ -4/5°;

^max 1757 «"1(C014).

Pound 0,67.14; H,9.50; 0,6.93; Br,17.l£ C27H4502Br requires 0,67.35; H,9.42; 0,6.65; Br,l6.6#

A small amount of pure (XIV) was refluxed in methanol until after about 30 minutes it was completely dissolved. Thin layer chromatography on silica gel/benzene: chloroform 1:1 and comparative nujol mull infrared spectroscopy showed that the (XIV) was quantitatively converted to (XII). A further small amount of pure (XIV) was dissolved in dioxan and a little lOfi sodium hydroxide solution added. Thin layer chromatography and comparative nujol mull infrared spectroscopy again showed that the (XIV) was completely converted to (XIII). 3-oxa-5a-cholestan-2-ol (XVIII). (XII) (2oo mg), lithium aluminium hydride (1.2g) and sodium dried ether (30 ml) were refluxed for 1 hour. The excess reagent was destroy­ ed by the cautious addition of ethyl acetate and the mixt­ ure then poured into ice cold 2N sulphuric acid (100 ml).

The ether layer was separated and the aqueous layer washed successively with ether then chloroform. The combined ether layer and washings were dried over anhydrous magnes­ ium sulphate and the solvent evaporated to leave a colour­ less solid residue. Recrystallisation successively frmm acetone, ethanol then petrol gave small needles (110 mg) - 69 - mp. 163.5-166°; [a] ^+15.5°.

Pound 0,80.26; H,11.99# C^H^gO^ requires 0,80.14; H,11.96?&

2-acetoxy-3-oxa-5a-cholestane (XIX). A mixture of (XVIII) (110 mg), pyridine (6 ml) and acetic anhydride

(4 ml) was heated on the steam "bath for 2 hours and set aside overnight at room temperature. Dilution with water precipitated the product which was extracted into ether.

The ether layer was washed successively with water, N sodium carbonate solution, water and dried over anhydrous magnesium sulphate. The ether was evaporated and the residue (36 mg) recrystallised from methanol to give plates mp.l05°; [a] D + 12.4° (2 dm tube); ^ x 1735 cm”1 (cci4).

Pound 0,77.71; H.11.25; Acetyl, 10.97® C _r,Hc_/^0_( mono acetate) 29 50 3 requires 0,77.97; H,11.28; Acetyl, 9.6%

Oxidation of (XVIII). Chromium trioxide (58 mg) in acetic acid (10 ml) was alowly added with cooling to

(XVIII) (100 mg) in acetic acid (5 ml). The mixture was set aside at room temperature for 80 minutes, ethanol (5 ml) added and the product precipitated by dilution with water. It was extracted into chloroform which was then washed with water, dried over anhydrous magnesium sulphate - 70 - and evaporated. The residue was recrystallised from methanol to give colourless plates mp.198-200°. The nujol mull infrared spectrum of this material was identic­ al with that of (XIII).

Pound 0,74.74; H, 11.12$

G27H46O3.CH^OH requires 0,74.66; H,11.10^

Chromous Chloride reduction of (XIV). Zinc .Amalgam: Zinc dust (80 g) was added with shaking to a solution of mercuric chloride (40 g) in 50^6 ethanol

(400 ml). The supernatent liquid was decanted and the finely divided solid material slurried out in water to leave convenient lumps (about pea size) of amalgam. The chromous chloride reagent was prepared by placing 20 ml of a N aqueous solution of chromic chloride, 5N with respect to hydrochloric acid, in a loosely stoppered burette together with several lumps of amalgam. Within 10 minutes the colour had changed from dark green to the brilliant sky blue of the chromous ion. This was very sensitive to atmospheric oxidation. A solution of (XIV)

(133 mg) in acetic acid (50 ml) was placed in a closed container having an inlet for nitrogen under the liquid surface and a gas outlet. The solution was purged with nitrogen for 15 minutes and the chromous chloride reagent

(5 ml) added against a positive flow of nitrogen. The reaction was warmed to 40°, allowed to cool and the con- - 71 - tainer sealed under nitrogen and set aside overnight at room temperature. Dilution with water (100 ml) pre­ cipitated the product which was extracted into chloroform. This extract was washed with water, dried over anhydrous magnesium sulphate and the solvent removed. The residue was recrystallised from methanol to give small needles

(80 mg) mp.199-200°.

4-hydroxy-3»4-seco-5a-cholestan-3-oic acid lactone (XX). Perhenzoic acid: Sodium (1 g) was dissolved in methanol

(20 ml) and the solution cooled in an ice/salt bath. Benzoyl peroxide (10 g) in chloroform (40 ml) was added slowly with shaking. The resulting sodium perbenzoate was extracted into ice cold water, the extract washed twice with cold chloroform and then acidified with ice cold N. sulphuric acid. The perbenzoic acid was extracted into 2x20 ml portions of chloroform to give a solution contain­ ing 0.0115 g perbenzoic acid per ml.

The perbenzoic acid reagent (31 ml) was added to cholestan-3-one (725 mg) in chloroform (2 ml) and set aside in the dark at room temperature for 46 hours. The solut­ ion was successively washed with sodium thiosulphate solution, water and evaporated to dryness. The residue was redissolved in ether and extracted twice with sodium carbonate solution, water and dried over anhydrous magnes­ ium sulphate. The solvent was removed and the residue - 72 -

recrystallised from methanol to yield needles (491 mg) mp.187-192°. Further recrystallisation from methanol gave the pure lactone as small needles mp.199-200°.

Found 0,80.66; H,11.63$

C27H46°2 re(luires 0,80.54; H,11.51$

The nujol mull infrared spectrum of this material was identical with that of the material obtained from the chromous chloride reduction of (XIV) and no depression of melting point w$s produced on admixture of these two com­ pounds . 2-(Bisphenyl carbinol)-3-oxa-5q-oholestane (XXI). Magnesium

turnings (8 g) , sodium dried ether (486 ml), bromobenzene

(33.5 ml) and a crystal of iodine were refluxed for 10 minutes to initiate the reaction which then proceeded spontaneously for a further half hour. The resulting dark brown cloudy solution was decanted from unreacted magnesium into a solution of (XII) (9 g) in sodium dried anisole (450 ml). The ether was distilled off and the remaining solution left on the bath for 6 hours and then

overnight at room temperature. The anisole was removed by steam distillation and the resulting light brown solid residue, which was shown by thin layer chromatography on

silica gel/chloroiorm to oe almost pure, crystallised from acetone to yield needles (9.86 g, 85$) mp.148-152°. - 73 -

Further recrystallisation from acetone gave needles which melted at 154-6°, resolidified into long filaments at

164-178° and these remelted at 184-186° [a] ^ -94.2°;

\>max 3566 om"1 ^ CCip •

Found: 0,84.09; H, 9.97; 0,6.10%

G39H56°2 reluires 0,84.15; H,10.14; 0,5.75%

Dehydration of (XXI) to the olefin (XXII). A mixture of

(XXI) (8.8 g) , acetic anhydride (88 ml) and acetic acid (550 ml) was refluxed for 7 hours, poured onto crushed ice and set aside overnight. The precipitated product was filtered off and recrystallised from acetone to give colourless rods (5.9 g, 70%) mp.151-153°. Further re­ crystallisation from acetone raised the melting point to

154-154.5°. [a]D -259.8°; \?max 1618 cm-1 (CCl^);

Amax 212mii( e=l.73^10^) ; 265mn( 6=1.54x10^)

Pound: 0,86.80; H.10.01; 0,3.95^

^39^54^ reluires 0,86.93; H,10.10; 0,2.96%.

3-oxa-5q-cholestan-2-one (XXXIII). A solution of (XXII)

(2 g) in dry ethyl acetate (100 ml) was cooled to about -40° in a dry ice/ethanol bath and ozonised for 50 minutes

By this time the solution was saturated with ozone and had assumed a light blue colour. It was allowed to come to room temperature, Adams catalyst (100 mg) added and the - 74 - solution hydrogenated at atmospheric pressure for 3 hours.

(180 ml H9 consumed) The catalyst was filtered off, the solution was concentrated to a small volume and water added. The mixture was boiled for 20 minutes, the remaining solv­ ent being allowed to steam distill off. The product was extracted into chloroform, the chloroform layer dried over anhydrous magnesium sulphate and evaporated to dryness. The residue was recrystallised fromirethanol to give colour­ less plates mp.135-137°. The mother liquors were reflux­ ed for 30 minutes in 3N alcoholic potassium hydroxide solution (10 ml), the solution diluted with water (50 ml) extracted with ether and then acidified with hydrochloric acid. The resulting precipitate was extracted into ether, the ether layer dried over anhydrous magnesium sulphate, and then evaporated to dryness. The residue was combined with the above crystalline material (1.2 g in all) and re­ crystallised repeatedly from methanol to give plates mp. 138-139°; [a]D +63°; \)max 1735 cm-1 ;CC14; .

Found: 0,80.09; H,11.24; 0,8.80#.

G26H440° recluires 0,80.35; H,11.4l; 0,8.23#

2,3-seco-5a-oholestane-2,3-dioic acid (XXIV). Gholestanol (50 g) in acetic acid (600 ml) was added to a mixture of potassium permanganate (75 g), water (100 ml) and acetic acid (500 ml). The mixture was heated on the water bath 75 - for 3.5 hours. Sufficient sodium bisulphite solution was added to decolourise the solution which was then diluted with water (1L). The product was extracted into chloro­ form and this extract washed with water and evaporated almost to dryness. . The residue was redissolved in ether and extracted with 13$ potassium hydroxide solution (150 rd) The aqueous layer was washed with ether, acidified with 5N hydrochloric acid and the resulting precipitate again extracted into ether. This extract was dried over anhydr­ ous magnesium sulphate and evaporated to dryness. The syrupy residue was recrystallised from petrol/benzene to yield colourless plates (11.3 g> 20$) mp.190-192°.

A-nor-5a-cholestan-2-one (XXV). 2,3-seco-cholestane- 2,3-dioic acid (XXIV) (11.3 g) and acetic anhydride (100 ml) were refluxed for 10 minutes. The anhydride was allowed to distill off slowly at atmospheric pressure until the temperature rose to 250°. The residue was then distilled under nitrogen at 5mm Hg and the distillate recrystallised successively from ethanol then methanol to yield almost colourless plates (5.47 g, 54$) mp.95-97.5°. Further re­ crystallisation from ethanol raised the melting point to

97-98°.

2-oxa-5a-cholestan-3-one (XXVI). A solution of tri- fluoroperacetic acid (0.008 moles approx.) in chloroform

(10 ml) was added with stirring to one of A-nor-cholest- - 76 - anone (1 g) in chloroform (15 ml). The mixture was stirred for 20 minutes and then refluxed for a further 1.5 hours. It was washed successively with water, dilute "bisulphite solution, water and then dried over anhydrous magnesium sulphate and evaporated to dryness. The residue was refluxed with alcoholic potassium hydroxide solution

(3N, 10 ml) for 1 hour, diluted with water (50 ml) and extracted with ether. The aqueous layer was acidified with 5N hydrochloric acid and the resulting precipitate extracted into ether. This extract was successively wash­ ed with N sodium carbonate solution, water and dried over anhydrous magnesium sulphate. The ether was evaporated and the residue (756 mg) recrystallised from methanol to give plates mp. 134-135°; [a] D + 20°; \)max 1736 cm”1 (001^). This material produced a sharp depression in melting point on admixture with (XXIII).

Pound: 0,79.81; H,11.19# 0,80.02; H,11.66#

0,79.53; H,11.21#

(Successive analyses were carried out after further re­

crystallisation from methanol).

G26H44°2 re(luires 0,80.35; H, 11.41#

C26H44°2,^GH 0H reQ-uires 0,79.49; H, 11.44# - 77 -

t-butyl hypochlorite: A solution of sodium hydroxide (80 g) in water (500 ml) was placed in a three necked flask fitted with stirrer and gas inlet. The flask was placed in a cold water bath and the contents cooled to

20°. t-butanol (96 ml) and water 600 ml were added and the mixture stirred to make an homogenous solution.

Chlorine gas was bubbled into the stirred cooled mixture for about 1 hour after which it was no longer absorbed.

The upper oily layer was separated, washed with 50 ml

portions of 10^S sodium carbonate solution until these were no longer acid to Congo red paper, then with 4x50 ml port­

ions of water and dried over calcium chloride. The t-butyl hypochlorite was kept in the dark in a refrigerat­ or.

2a-Chloro-5a-Cholestan-3-one. A solution of cholestanol

(5 g) in acetic acid (150 ml) at 65° was treated rapidly with t-butyl hypochlorite (3*08 ml; 2.1 mole) and the temperature held at 70° for 2 hours. On cooling to

slightly below room temperature the product separated as needles (2.48 g; 45$) mp.173-177°. This was shown by thin layer chromatography to be sufficiently pure for the next

step.

2a-chloro-4-hydroxy-3,4-seco-5a--cholestan-3-oic acid

lactone (XXX). A vigorously stirred mixture of 2a-

chloro-cholestan-3-ona (2 g) and disodium hydrogen phosph- - 78 - ate (5 g) in chloroform (26 ml) was treated with a solut­ ion of trifluoroperacetic acid (0.01 moles approx.) in chloroform (8 ml). The mixture was refluxed for 2 hours and then left overnight at room temperature. It was successively washed with water, sodium bisulphite solut­ ion, water and dried over anhydrous sodium sulphate. The solvent was evaporated under reduced pressure and the yellow solid residue recrystallised from acetone to give needles (1.32 g; 49$) mp.186-193°. Further recrystallis­ ation from acetone and methanol/chloroform gave colourless rods mp. 194-5°; [a] D -4.3°; \)max 1762 cm-1 (CC14).

Found C,74.07; H,10.48$.

C27H45°2 01 re<3Mires 0,74.19; H,10.38?£.

A little of the pure material (XXX) was dissolved in dio- xan and a few drops of 3N sodium carbonate solution added. The solution was acidified and the organic material com­ pletely reprecipitated by dilution with water. This was shown by nujol mull infrared comparison and mixed melting point to be identical with the carboxylic acid (XIII).

2a-iodo-5a-oholestan-3-one. 2ct-bromo cholestan-3-one

(3 g) and sodium iodide (3-6 g) in acetone (75 ml) were refluxed for five and a half hours. Sufficient sodium thiosulphate solution was added to discharge the faint iodine colour and the solution cooled to deposit a floc- culent white precipitate. A yield of 3.2 g colourless - 79 -

needles mp.134-139° was obtained and this material was shown by thin layer chromatography on silica gel to be quite pure.

5a-Cholest-l-en-3-one. k vigorously stirred mixture of

iodocholestanone (2 g) and disodium hydrogen phosphate (5 g) in chloroform (26 ml) was treated with a solution of trifluoroperacetic acid (0.01 mole approx.) in chloro­ form (8 ml). An immediate strong violet colour was prod­

uced. The mixture was refluxed for 2 hours and then successively washed with water, dilute bisulphite solution

(until colourless), water and dried over anhydrous sodium

sulphate. The chloroform was removed under reduced press­ ure and the residue lecrystallised from acetone to give

colourless crystals (0.30g) mp.95-98°. The mother liquors

(1.31g brown glass) were dissolved in hexane and chromato­ graphed on silica gel (60g). Elution with hexane+20$ benzene gave 0.47 g crystalline material shown by thin layer chromatography to be identical with that previously

isolated. ho further crystalline material could be eluted. Recrystallisation from methanol gave large glistening plates mp.98-99°; ^mo„230m^,(log e=4) fc]-n+59.4°.

(Elsevier mp.96-100°; [a]D+57.5°; ^max^BOmiijClog e-4) •

The above experiment was repeated on a test tube scale by

dissolving a little iodocholestanone in chloroform and

adding the trifluoroperacetic acid reagent. The violet - 80 - colour was again produced immediately. The addition of 87% hydrogen peroxide, 40% peracetic acid and 87% hydrogen peroxide plus p-toluene sulphonic acid respectively to separate solutionsof iodo-cholestanone in chloroform did not produce the violet colour.

The extraction of Friedelin and Gerin. Commercial gran­ ulated cork (2360 g) was Soxhlet extracted with chloro­ form in 700 g lots. Each lot was extracted for about 20 hours. Upon evaporation of the combined extracts to a small volume (c.a. 600 ml) crude crystalline cerin (1.8 g) was deposited. This was collected and the mother liquor evaporated to a syrup, treated with boiling ethanol (II) and filtered while hot to leave crude solid friedelin. This was dissolved in benzene and filtered through a col­ umn of alumina (200 g). The friedelin so obtained (8.1 g) was dissolved in 1400 ml of petrol containing 30% benzene and carefully chromatographed on alumina (350 g), eluting with this solvent. The middle fractions were combined

(6.74 g) and recrystallised from chloroform/acetone to give needles mp.260.5-263°; [aJD-21°. Thin layer chroma- tography on silica gel/chloroform showed this material to be of high purity. The cerin was recrystallised from chloroform to give needles mp. 261-263° [a] p-50.2°.

2a-bromofriedelin. Friedelin (1 g) in chloroform (50 ml) was treated with 45% hydrogen bromide in acetic acid (1 ml) -81- and stirred for 10 minutes. A solution of bromine in chloroform (1.1 g/lO ml; 3*9 nil) was added dropwise with stirring during 10 minutes. The solvent was evaporated under reduced pressure to a small volume and methanol added to precipitate the product as pale yellow needles

(1.025 g) mp.198.210°dec. Recrystallisation from methanol gave colourless needles (682 mg) mp.210° dec. [a]^-138°

Attempted Baeyer-Villiger oxidation of bromofriedelin. A mixture of bromofriedelin (100 mg), disodium hydrogen phosphate (2 g) and trifluoroperacetic acid (0.005 moles approx.) in chloroform (10 ml) was refluxed and aliquots removed after 2,4,6 and 10 hours respectively and examined by thin layer chromatography (silica gel/benzene). All these aliquots showed that starting material only was present in appreciable amount. The reaction was repeated in the absence of the disodium hydrogen phosphate. An aliquot removed after 2 hours refluxing was shown by thin layer chromatography to contain a highly polar compound and no starting material. The reaction mixture was successively washed with dilute sodium bisulphite solution, water, and dried over anhydrous sodium sulphate. The chloroform was evaporated to leave a yellow oily residue, which could not be crystallised. Since this material could also not be satisfactorially chromatographed on thin layers of silica gel, column chromatography was not - 82 -

attempted. Its infrared spectrum showed strongly hy­ drogen bonded hydroxyl absorption and carbonyl bands at 1720 and 1790 cm~1 (Infracord). The above reactions were again repeated i.e. in the presence and absence, respectively, of the disodium hy­ drogen phosphate buffer. The reaction mixtures, however, were not refluxed but were left to stand in the dark at

room temperature and aliquots examined chromatographically at intervals. After one day the bromofriedelin in the reaction containing no buffer was completely converted to

an intractible oil similar to that described above, where­

as after 3 weeks the buffered reaction mixture contained mainly starting material together with a little highly polar material. Baeyer-Villiger oxidation of Gerin.in Chloroform solution: Gerin (2 g) was refluxed in chloroform (230 ml) with 40$ peracetic acid (23 ml) for 12 hours. The mixture was left overnight at room temperature, washed with sodium bisulphite solution, water, and dried over anhydrous mag­ nesium sulphate. The solvent was evaporated under reduced pressure and the residue redissolved in ether. Extract­ ion with 3N sodium carbonate solution gave an acidificat­ ion a colourless precipitate (1 g) which was recrystallised from ethyl acetate to give large rods mp.287-288°, identified as friedelin dicarboxylic acid (XXXVIII) by - 83 - mixed melting point and nujol mull infrared spectrum comparison with an authentic sample. The ether layer was dried over anhydrous magnesium sulphate and the ether removed to leave a colourless solid residue (0.59 g). Recrystallisation of this material from chloroform/methan- ol gave 2,3-dihydroxy-3,4-seco-friedelan-4-oic acid lactone (XXXTV, R=H) as small needles mp. 318-324°;

La] un +16.3°; max 2317,1706 cm”1 (halocarbon oil mull;.

Pound 0,78.51; H,11.07$

G30H50°3 requires 0,78.55; H, 10.99#

In buffered acetic acid solution: A mixture of cerin (2 g) and anhydrous sodium acetate (40 g) in acetic acid (1300 ml) was treated with 40^& peracetic acid (40 ml).

The mixture was set aside in the dark at room temperature for 14 days. Dilution with water (31) precipitated the product which was extracted into chloroform. Proceeding as previously the chloroform layer was washed, dried, evaporated to dryness and the residue redissolved in ether,

Sodium carbonate extraction yielded friedelin dicarboxylic acid (XXVIII) (1.22 g) while the remaining ether layer yielded a yellow glass (0.53 g). Thin layer chromatography showed this to be a complex mixture. Repeated recrystall­ isation from methanol gave a low yield of crystalline material mp.270-273° identified as impure (XXXIV, R=H) by - 84 - its nujol mull infrared spectrum.

Acetyl Qerin. Cerin (2.4 g) pyridine (72 ml) and acetic anhydride (48 ml) were heated together on the water hath for 2 hours. The mixture was poured into water (1L) and set aside at room temperature overnight. The colourless precipitate of cerin acetate was filtered off and recrystal­ lised from ethanol/chloroform and ethyl acetate-chloroform to give needles mp.255-258°. [a] ^ -31°.

2(3-acetoxy-3-hydroxy-3,4-seco-friedelan-4-oic acid lactone (XXXIV, R=Ac). A vigorously stirred mixture of Cerin acetate (0.98 g), anhydrous disodium hydrogen phosphate (1 g) in methylene chloride (30 ml) was treated dropwise with a solution of trifluoroperacetic acid (approx. 0.005 mole) in methylene chloride (4 ml). Stirring was continued at room temperature for 2 hours. The insoluble salts were filtered off and washed with a little methylene chloride.

The combined filtrate and washings were then washed successively with sodium bisulphite solution, water and dried over anhydrous magnesium sulphate. Evaporation of the solvent gave a colourless solid residue (1 g) which on recrystallisation from ethanol gave needles (237 nig), mp. 206.208°. Itirther re crystallisation from methanol gave pure (XXXIV, R=Ac) mp.223.5-228°; tn] D+14.8° (2 dm.tube) ;

0raax 1748,1740 cm_1(CCl4). - 85 -

0,75.80; Pound: H,10.28^ 0,75.89; H,10.75$

G32H52°4*^C5H3OH re(luires 0,75.60, H,10.47£

Hydrolysis of this material in N/2 alcoholic alkali follow­

ed by acidification gave a compound identified as (XXXIV, R=H) by melting point, mixed melting point and nujol mull infrared spectrum comparison.

The mother liquors yielded no further crystalline material. They were evaporated to dryness and the residue refluxed in for 1 hour in R alcoholic alkali. Acidificat­ ion and dilution with water precipitated a product which was extracted into ether. This extract was dried over anhydrous magnesium sulphate, the ether evaporated and the residue recrystallised from chloroform/methanol to yield

colourless needles (476 mg) mp.318-324°, identified as

(XXXIV, R=H) by mixed melting point and nujol mull infrared spectrum comparison.

Acetylation of (XXXIV, R=H). (XXXIV, R=H) (80 mg) was re­ fluxed in acetic anhydride (10 ml) for 2 hours and left overnight at room temperature. The mixture was poured into water, set aside at room temperature during four hours and the precipitated product collected (83 mg). Re­ crystallisation from methanol gave needles mp.223.5-228° shown to be identical with (XXXIV, R=H) by mixed melting point and nujol mull infrared spectrum comparison. ACKNOWLEDGMENTS

The author would like to express his gratitude to Dr. J.L. Courtney for his super­ vision and valuable guidance throughout the course of this work.

The author is indebted to Dr.E.Challen for the analytical figures quoted herein. - 87 -

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