Synthesis of deuterated methyl 6,9,12-octadecatrienoate geometric isomers

Chemistry and Physics o f Lipids, 35 (1984) 117-125 Elsevier Scientific Publishers Ireland Ltd.

SYNTHESIS OF DEUTERATED GEOMETRIC ISOMERS

METHYL

117

6,9,12.OCTADECATRIENOATE

HENRY RAKOFF Northern Regional Research Center, Agricultural Research Service, U.S.. Department of Agri. culture, Peoria, IL 61604/U.S.A.) Received March 6th, 1984 accepted April 18th, 1984

revision received April 16th, 1984

Methyl cis-6,cis-9,cis-12-octadecatrienoate-15,15,16,16.d 4 and the corresponding eis.cis, trans isomer were obtained by coupling hexyl-d~-triphenylphosphonium bromide and methyl 12-oxo-cis-6,cis-9-dodecadienoate by the Wittig reaction. The deuterated phosphonium salt was prepared from 3-hexynol by catalytic deuteration of the corresponding tetrahydropyranyl ether and intermediate formation of the bromide. The dienoic aldehyde ester was obtained through the intermediate dioxanyl and dimethoxy derivatives from the Wittig coupling of methyl 9-oxo-cis-6-nonenoate with [2-(l,3-dioxan-2-yl)ethyl]-triphenylphosphonium bromide. The monoenoic aldehyde ester was prepared in a similar manner by the Wittig reaction between methyl 6-oxohexanoate and the dioxanylphosphonium salt. The saturated aldehyde ester was obtained, through several steps, from the ozonolysis of cyclohexene. Geometric isomers formed during each of the Wittig reactions were separated by silver resin chromatography. 13C Nuclear magnetic resonance chemical shifts for the compounds prepared are presented. Keywords. synthesis; methyl 6,9,12-octadecatrienoates-d,; I~-NMR; cyclic acetal esters: dimethyl acetal esters; aldehyde esters; deuterated 3,-linolenates; geometric isomers. Inlroduction We have been involved, for several years, in a study o f the metabolism in humans o f isomeric fats formed during the hydrogenation o f soybean oil [1,2]. For this purpose, we have prepared deuterated analogs of many o f these monoenoic and dienoic acids [3]. We have also prepared all the geometric isomers of methyl 9,12,15octadecatrienoate [4], a member of the c~-3 acid series. The current interest in ~ - 3 and co-6 acids prompted us to devise a synthetic scheme for the preparation of deuterated analogs o f methyl 6,9,12-octadecatrienoate, a member of the co-6 series. Methyl cis-6,cis-9,cis.12-octadecatrienoate, methyl-3,-linolenate, was identified in the seed oil o f the evening primrose, Oenothera biennis, family Onagraceae, by Riley [5] in 1949 and has been found in several other families [6]. The all cis isomer was synthesized by Osbond et al. [7,8] from 1-bromo-2-octyne via acetylenic coupling and Lindlar reduction. Stoffel [9,10] prepared the tritiated analog labeled in the 17,17,18,18-positions and the 1-14C analog by similar techniques. Klok et al. 0009-3084/84/$03.00 © 1984 Elsevier Scientific Publishers lreland Ltd. Published and Printed in Ireland

118

[11] prepared the 6.trans,9-cis, 12.cis isomer from 1,4-dichloro.trans.2.butene via acetylenic coupling, malonic ester chain extension and Lindlar reduction. The mass spectrum [12] and ~3C-NMR chemical shifts [ 1 3 - 1 5 ] for the all cis isomer have been recorded. Each of the syntheses referred to can be used to prepare only one of the eight possible geometric isomers of methyl 6,9,12-octadecatrienoate. This paper describes a general synthetic method that can be used to prepare any or all of these geometric isomers and its application to the synthesis of the cis,cis, cis and the cis, cis, trans isomers. In our synthesis, methyl 6-oxohexanoate, 1, is converted via two Wittig

OCH(CH2)4COOCH3 + QCH2CH2P(C6Hs)3Br C4HeLi THF

1

2

QCH2CH=CH(CH2)4COOCH 3 p-CH3C6H4SO3H CH3OH, .4 v

3 (85%)

CH3CN-H20 Co~. HCl

(CH30) 2CHCH2CH--CH (CH2)4COOCH3

4 (90%) OHCCH2CH~--CH(CH2)4COOCH3

5 (95%) C4HgLi • QCH2CH=CHCH2CH~CH(CH2)4COOCH3 THF

5+2

6 (53%) p-CH3C6H4SO3H CH3OH, .4

(CH30) 2CHCH2CH--CHCH2CH= C H (CH2)4COOCH3

7 (93%) CH3CN-H20 Con.HCI :- OHCCH2CH=CHCH2CH=CH(CH2)4COOCH3

8 (62%) 8 + CH3CH2CD2CD2CH2CH2P(C6Hs)3Br C4HgLi THF

9

CH3CH2CD2CD2CH2CH~ CHCH2CH= CHCH2CH---CH (CH 2)4COOCH3

10 (59%)

CH~--O * Q is CH2

\

/

H-

CH2--O

Reaction Scheme

119 couplings with [2-(1,3-dioxan-2.yl)ethyl]triphenylphosphonium bromide, 2, to methyl 12-oxo-6,9-dodecadienoate, 8. After each double bond is generated, ~he geometric isomers and conjugated isomers formed are separated by silver resin chromatography. The diunsaturated aldehyde ester, 8, is then coupled in a Wittig reaction with hexyl-d4-triphenylphosphonium bromide, 9, to yield methyl 6,9.12-octadecatrienoate- 15,15,16,16~d4 geometric isomers, 10, as shown in the reaction scheme.

Experimental Reagents [2-(l,3-Dioxan-2-yl)ethyl] triphenylphosphonium bromide may be prepared from acrolein [16] or purchased, as may cyclohexene and butyl lithium, from Aldrich Chemical Company. Hexyl-d4-triphenylphosphonium bromide was prepared t'rom 3-hexynol as previously described [17].

Procedures Silver resin chromatography The macroreticular resin used for the separations was Rohm and Haas XNI010 sulfonic acid resin ground to the mesh size indicated in the synthetic descriptions. For the Ag/H columns, the indicated percentage of hydrogen ions was displaced by Ag ions [18]. For the Ag/Na columns, the hydrogen ions were displaced by sodium ions and then the indicated percentage of sodium ions was displaced by silver ions [ 19]. Eluant was methanol, unless otherwise indicated.

Gas chromatography A 50 meter × 0.25 mm OV275 WCOT capillary GC column was used for analyzing binary mixtures of geometric isomers. For other analyses, a 6 ft X 4 mm column packed with 3% EGSSX on 100/120 Gas Chrom Q was employed.

'3C.NMR 13C-NMR spectra were recorded with a Bruker WM 300 WB pulsed Fourier transform spectrometer operating at 75.5 MHz. Typically, 2500 transients were collected from solutions in CDCI3, which served as both the internal lock and secondary reference, using 5 mm tubes. Sweep widths of 200 ppm and 8K real data points limited acquisition time to 0.54 s and were used to obtain chemical shift values to within -+1.85 Hz, i.e. -+0.05 ppm. A pulse width of 3 ps (40 °) was employed with no delay between pulses. Decoupling power was held to approx. 1 W to provide adequate broadband decoupling power while minimizing sample heating.

Mass spectroscopy Mass spectra were determined on a Nuclide 12-90-DF spectrometer with 70 eV ionizing energy.

120

Syntheses Preparation of dioxanyl esters Methyl 8-dioxanyl-6-octenoate, 3 Dioxanylethyltriphenylphosphonium bromide, 2 (50.6 g, 110.7 mmol), was slurried in tetrahydrofuran (250 ml) in a 1-1 3-necked flask equipped with a mechanical stirrer, a low temperature thermometer, and a N2 inlet and outlet and cooled in an ice-salt bath to 0°C. Butyl lithium (1.5 M in hexane, 80 ml, 120 mmol) was added in portions over 7 min with the temperature not exceeding 11°C. One hour later, methyl 6-oxohexanoate (16 g, 111 mmol) dissolved in THF (5 ml) was added to the brown liquid over 3 min. The temperature rose to 20°C and the color became much lighter. Fifteeen minutes later the ice bath was removed and 45 min later the reaction mixture was shaken with saturated NaCI (100 ml)and extracted into ether (2 × 50 ml). The ether layer was dried (Na2SO,). After removal of the drying agent and solvent, the mixture was distilled to give 3 (23.17 g, 86% yield) b.p. 90-110°C/0.1 torr. Purity by capillary GC on OV275 was 97% and the trans/ cis ratio was 31 : 69.

Methyl l l-dioxanyi-6,9-undecadienoate, 6 In a similar manner, 2 and 5 were combined with butyl lithium in THF to give 6 (6.47 g, 53% yield) b.p. 130-155°C/0.1 Tort. Capi0.ary GC analysis indicated 82% of the 6,9-isomer (15% ct, 85% cc) and about 12% conjugated isomers.

Preparation of dimethoxy esters Methyl 12,12-dimethoxy-6, 9-dodecadienoate, 7 Methyl l l-dioxanyl-6,9-undecadienoate, 6, (5.02 g, 17.8 mmol), methanol (700 ml) and p-toluenesulfonic acid (0.4 g) were heated at reflux in a 1-1, roundbottomed flask for 30 min. The reaction mixture was cooled, stirred with solid Na2CO3 and filtered. The methanol was removed on the rotary evaporator, and the solid remaining on the walls was extracted into ether (2 X 25 ml). The cloudy ether solution was washed with H20 (1 × 10 ml) and the clear ether solution resulting was dried (Na2SO4). Removal of the drying agent and solvent left a liquid (4.48 g, 93% yield) which, on capillary GC on OV275, showed about 85% non-conjugated acetal ester, 7 (ct/cc, 19:81), about 1 l% conjugated isomers and about 3%unreacted 6. The reaction may be repeated if desired to convert the remaining dioxanyl ester to the dimethoxy ester.

Methyl 9, 9-dimethoxy-6-nonenoate, 4 In a similar manner, compound 4 was obtained from compound 3. After one reaction, there was about 7% unreacted starting material and about 4.5% conjugation. Repetition of this reaction gave compound 4 in about 90% crude yield, containing

121 about 1.5% unreacted starting material and about 6% conjugation. To prevent further isomerization, this material was not distilled but was used immediately in the Wittig reaction.

Preparation of aldehyde esters Methyl 6-oxohexanoate, 1 Methyl 6,6-dimethoxyhexanoate (48.4 g, 255 mmol) was dissolved in water ~65 ml) and acetonitrile (165 ml) in a 500-ml, round-bottomed flask equipped with a magnetic stirrer and an inlet and outlet for maintaining a nitrogen atmosphere [20]. Concentrated hydrochloric acid (30 drops) was added to the solution, and 4.5 h later the reaction mixture was concentrated to 60 ml (2 phases) on the rotary evaporator. The mixture was extracted with ethyl ether (2 X25 ml) and dried (Na2SO4). Removal of the drying agent and solvent gave a liquid which was distilled (b.p. 60-700C/0.15-0.30 Torr) to give methyl 6-oxohexanoate (33.60 g, 91% yield). GC on OV275 shows the product to be better than 98% pure.

Methyl 9-oxo-6-nonenoate, 5 Compound 5 was prepared from 4 in a similar manner, with a reaction time of about 18 h, in about 95% crude yield. It contained about 17% conjugated isomers. When the reaction was run in aqueous acetic acid [21], the product contained about 12% conjugated isomers.

Methyl 12-oxo-6, 9-dodecadienoate, 8 Compound 8 was prepared from 7 in a similar manner in about 61% crude yield. It contained about 38% conjugated isomers. When the reaction was run in aqueous acetic acid [21 ], the product contained about 22% conjugated isomers. Neither 5 nor 8 was distilled to obviate further isomerization but was used immediately in the Wittig reaction.

Methyl 6, 9,12-octadecatrienoate-15,15,16,16-d4 isomers, 10 Hexyl-d,rtriphenylphosphonium bromide, 9, (2.16 g, 5 mmol) was slurried in tetrahydrofuran (25 ml) in a lO0-ml, 3-necked flask equipped with a mechanical stirrer, a low temperature thermometer and a nitrogen inlet and outlet and cooled in an ice-salt bath to 2°C. Butyl lithium (1.5 M in hexane, 6 ml, 9 mmol) was added and an orange color developed. The ice bath was removed and the reaction mixture was stirred for 35 min. It was then cooled in a dry ice.isopropyi alcohol bath to -55°C and methyl 12~xo,cis.6,cis.9-dodecadienoate, 8, (1 g, 4.46 mmol) dissolved in tetrahydrofuran (2 ml) was added. The mixture became thicker and lighter in color. After 30 rain, a sample was withdrawn, added to saturated NaC1 solution, dried and analyzed by capillary GC. About 25% of the area was represented by two conjugated isomers in a ratio of 73 : 27. For the non-conjugated isomers, the ratio of cct/ccc was approx. 8:92. Later (2.5 h), methanol (7 ml) was added to the

122 mixture at -52°C to give a clear yellow solution. The bath was packed with dry ice and left to warm up slowly overnight. The next morning, the clear red liquid was washed with saturated NaC1 solution (10 ml) and the red upper layer was then dried (Na~SO4). The drying agent and solvent were removed and the product was distilled in vacuo. A product mixture was obtained (0.62 g, 50% yield), b.p. 132-148°C/ 0.08 Torr that analyzed by capillary GC as 22% conjugated isomers (ratios 33 : 66) and 76% non-conjugated isomers (ratio cct/ccc, 63 : 37). This mixture was separated on a 62% Ag/Na XN1010 column (80/120 mesh) with methanol as eluant. Mass spectroscopic analyses showed 88% d4, 4.4% d2, 3.5% d3 and 1.1% d6. Average number of deuterium atoms per molecule is 3.87. Results mad Discussion Methyl 6-oxohexanoate, 1, was obtained from the ozonolysis of.cyclohexene by the method previously described [17] for the preparation of methyl 12-oxododecanoate from cyclododocene. Reaction of compound 1 with compound 2 with butyl lithium in tetrahydrofuran gave a mixture of the cis (70-75%) and trans (30-25%) isomers of 3, methyl 8-dioxanyl-6-octenoate. This mixture was separated into the individual isomers on a 100% Ag/Na column [18]. If compound 3 is passed through a 100% Ag/H column in methanol, partial transacetalation occurs on the column (which contains some free sulfonic acid groups) to form the isomers of 4, methyl 9,9-dimethoxy-6-nonenoate, and a clean separation of the isomers is not realized. Compound 3 is conveniently converted to compound 4 with methanol and p-toluenesulfonic acid, and the geometric isomers of 4 may be separated equally easily on a 100% Ag/Na or a 100% Ag/H column. Selective hydrolysis [20] of the cis isomer of 4 with concentrated HCI gave 5, methyl 9-oxo-cis-6.nonenoate containing about 17% conjugated isomers. StoweU and Keith [21] claim that hydrolysis of their/3,3,-unsaturated dimethyl acetals with aqueous acetic acid gave only the /3,3,-unsaturated aldehyde uncontaminated with the 0~,/3 isomer. With aqueous acetic acid hydrolysis of 4, we were able to obtain 5 containing only about 12% conjugated isomers. Reaction of the cis isomer of 5 with compound 2 yielded 6, the cis, cis and cis, trans isomers of methyl l l-dioxanyl-6,9-undecadienoate. Transacetalation of 6 with methanol and p-toluenesulfonic acid gave the cc and ct isomers of 8, methyl 12,12-dimethoxy.6,9-dodecadienoate. The geometric isomers were separated from each other and from the conjugated isomers that were formed during the hydrolysis of 4 to S by silver resin chromatography on a 91% Ag/H column. Methyl 12-oxocis-6,cis-9-dodecadienoate, 8, was obtained by hydrolysis of the cis,cis isomer of 7, together with 38% conjugated isomers when HCI in aqueous acetonitrile was used and 22% conjugated isomers when aqueous acetic acid was used for the hydrolysis. Aldehyde ester 8 was caused to react with hexyld4-triphenylphosphonium bromide, 9, at ice bath temperature to get predominant (90%) formation of the cis, cis, cis isomer, or at -50°C under conditions to effect thermodynamic control [22] of the product mixture to get predominant (63%) formation of the cis, cis, trans isomer.

123 From each reaction mixture, the ccc and cct isomers were separated from each other and from the conjugated isomers formed during the preparation of 8 by partial silver resin chromatography on a 62% Ag/Na column. The all cis isomer had the same retention time on the OV275 capillary column as the naturally occurring all cis isomer in Borago officinalis methyl esters [23]. Table I lists the chemical shifts of the various intermediates prepared. The values listed are consistent with the structures and configurations assigned. The chemical shift o f a carbon alpha to a trans double bond is always approx. 5 ppm further downfield than that o f a carbon alpha to a cis double bond as expected [24,25]. The values for a methylene group between a double bond and a dioxanyl group are 33.40 (cis) and 38.60 (trans), between a double bond and a dimethyl acetal are 31.10 (cis) and 36.10 (trans), and between a double bond and a carbonyl group much further downfield at 42.60 (cis). Table II lists the chemical shifts for the isomeric 6,9,12-octadecatrienoates-d4 prepared and for the naturally occurring all cis isomer. Our results for the all cis isomer compare very well with the values recorded in the literature [15]. Our compound differs from the natural compound in that it contains deuterium atoms on C-15 and C-16. The signal from carbons bearing two deuterium atoms is diminished to such an extent that it is usually not detected. We did not obtain a chemical shift of 31.6 in either of our compounds for C-16. We did obtain a peak at 29.2, but this

TABLE 1 13C-NMR CHEMICAL SHIFTS OF INTERMEDIATES a Compound no. 1 Carbon b

5 6 7 8 9 I0

11 12

3 6c

43.30 201.50

26.90 123.50 131.50 33.40 101.90 _c

4

5

~ 6t

32.00 124.30 132.80 38.60 I01.90 _¢

27.00 124.00 131.70 31.15 104.30 d

27.30 I18.70 134.70 42.60 199.20

6

7

8

6c,9c

6c,9t

6c,9c

6c,9t

26.90 129.75 128.10 25.90 123.45 130.55

26.80 130.00 127.90 30.60 124.50 131.50

26.80 129.75 127.95 25.80 123.75 130.50

26.60 129.85 127.65 30.40 124.70 131.30

33.60 38.80 I01.90 101.90 c c

31.10 36.10 104.30 104.30 d d

26.90 130.30 127.10 25.60 I18.55 133.30

42.50 199.20

.appm downfield from (CH~)4Si. bChemical shifts are 173.90, 33.90, 24.60 and 29.00 for carbons 1-4 and 51.30 for the ester methyl carbon. For compound no, 1, carbon 4 is 21.40. CChemical shifts are 66.90 and 25.90 for carbons a and ~, respectively, to the oxygens in the dioxanyl group. dChemical shift is 52.90 for the acetal methyl carbon.

124 TABLE II ~C-NMR CHEMICAL SHIFTS FOR METHYL 6,9,12-18 : 3-15,15,16,16-d 4 ISOMERS" 6c,9c,12c

6c,9c, 12t

6c,9c, 12c

6c,9c, 12t

Carbon

Lit. [ 15 ]

Obs.

Obs.

Carbon

Lit. [15]

Obs.

Obs.

1 2 3 4 5 6 7 8 9

173.83 33.99 24.70 29.23 26.96 129.60 128.38 25.74 128.14

174.00 34.10 24.72 29.23 26.97 129.68 128.37 25.77 128.17

174.05 34.09 24.71 29.22 26.95 129.60 128.42 b 25.69 128.25b

10 11 12 13 14 15 16 17 18 OCH~

128.46 25.74 127.74 130.42 27.33 29.44 31.64 22.66 14.07 51.31

128.46 25.77 127.70 130.54 27.11

128.42b 30.54 128.02 131.15 32.36

22.40 14.05 51.45

22.36 14.03 51.43

appm down field from (CH3hSi. tCl'entativeassignment. is assigned to C-4. Tulloch [26] has studied the effect of gem dideutero, CD2, carbons on the chemical shifts of adjacent carbons. He found that the shift for the carbons alpha to a CD2 group is about 0.2 ppm less than for a carbon alpha to a CH2 group. We note this difference in both C-17 and C-14 of the all cis isomer. The chemical shifts for C-5, C-8, C-11 and C-14 are what would be expected [24, 25] for carbons alpha to one or two double bonds in both compounds. These shifts, together with the absence of the shift at 31.6 for C-16 and the isotope effects on C-14 and C-17, confirm the structure and configurations assigned on the basis of the synthetic scheme. Acknowledgements We thank D. Weisleder and L.W. Tjarks for NMR spectroscopy and E.A. Emken for helpful discussions.

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125

8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26

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