Evaluation of lipase from Candida rugosa in the resolution of endo- (±)- 1,8,9,10,11,11-hexachloropentacyclo- [6.2.1.13,6.02,7.05,9]dodecan-4-alcohol

Article abstract of DOI:10.1016/S0957-4166(98)00259-6

endo-(±)-1,8,9,10,11,11- Hexachloropentacyclo[6.2.1.1^3,6.0^2,7.0^5,9)]dodecan-4-ol (±)-7 and exo-(±)1,8,9,10,11,11- hexachloropentacyclo[6.2.1.1^3,6,0^2,7.0^5,9]dodecan-4-ol (±)-4 have been prepared and the enantiomeric enrichment capacity of the lipase from Candida rugosa in the transesterification with vinyl acetate of these compounds was evaluated. It was verified that the lipase recognize only the alcohol (±)-7, producing endo-(+)- 1,8,9,10,11,11- hexachloropentacyclo[6.2.1.1^3,6.0^2,7.0^5,9]dodecan-4-yl acetate (+)- 8 with ee >95% and conversion of 44% as the only product.

Full text of DOI:10.1016/S0957-4166(98)00259-6

TETRAHEDRON:
ASYMMETRY
Pergamon
Tetrahedron: Asymmetry 9 (1998) 2579–2585
Evaluation of lipase from Candida rugosa in the resolution of
endo-(ꢀ)-1,8,9,10,11,11-hexachloropentacyclo-
[6.2.1.13,6.02,7.05,9]dodecan-4-alcohol
Valentim E. U. Costa,∗ João Alifantes and Jose E. D. Martins
Instituto de Química, Universidade Federal do Rio Grande do Sul, Av. Bento Gonçalves, 9500, 91501-970 Porto Alegre-RS,
Brazil
Received 17 April 1998; accepted 24 June 1998
Abstract
endo-(ꢀ)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6.02,7.05,9]dodecan-4-ol
(ꢀ)-7
and
exo-(ꢀ)-
1,8,9,10,11,11-hexachloropentacyclo[6.2.1.13,6.02,7.05,9]dodecan-4-ol (ꢀ)-4 have been prepared and the
enantiomeric enrichment capacity of the lipase from Candida rugosa in the transesterification with vinyl acetate
of these compounds was evaluated. It was verified that the lipase recognize only the alcohol (ꢀ)-7, producing
endo-(+)-1,8,9,10,11,11-hexachloropentacyclo[6.2.1.13,6.02,7.05,9]dodecan-4-yl acetate (+)-8 with ee >95% and
conversion of 44% as the only product. © 1998 Elsevier Science Ltd. All rights reserved.
1. Introduction
The synthesis of hexachlorinated polycyclic systems, the insecticides isodrin, endrin and its derivatives
have been reported in the early 1950s.1 Soloway et al.2 verified that chlorinated compounds with the
1,4,5,8-dimethanonaphthalene nucleus undergo unusual skeletal rearrangements in their reactions. Win-
stein et al.3 observed Wagner–Meerwein rearrangements in solvolysis reactions of polycyclic derivatives,
whose mechanisms involve the formation of non-classical ions. The degradation of chlorinated polycyclic
systems, usually involving hydrolytic, oxidative and intramolecular rearrangement reactions, leaves the
basic carbon skeletal system intact.4
The investigation of rigid cyclic systems with unique conformations have contributed to the under-
standing of several aspects of NMR spectroscopy in relation to geometry and steric parameters. Long-
range steric effects of tetracyclododecanes endo–endo and endo–exo have been studied in our group by
means of 13C and 1H NMR.5–7
∗
Corresponding author. E-mail: valentim@if.ufrgs.br
0957-4166/98/$ - see front matter © 1998 Elsevier Science Ltd. All rights reserved.
PII: S0957-4166(98)00259-6

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Although some of these structures possess chirality, these studies were carried out with racemic
compounds. Efficient methodologies for enantiomeric analysis and resolution of these compounds are
lacking. Recently our work group has demonstrated that chiral and achiral chemical shift reagents can be
good analytical alternatives for determination of enantiomeric purity.8
In this paper, we describe the preparation of the hexachlorinated endo- and exo-pentacyclododecanols
(ꢀ)-4 and (ꢀ)-7, their transesterification reaction with vinyl acetate catalyzed by lipase from Candida
rugosa and the resolution of endo-(+)-1,8,9,10,11,11-hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-
4-yl acetate (+)-8 with high optical purity.
2. Results and discussion
The racemic alcohol (ꢀ)-4 was obtained as shown in Scheme 1. Following the
modified Lidov2 method, the isodrin
1 is completely consumed in 5 min by treat-
ment with acetic acid and catalytic sulfuric acid, giving
a
mixture of four prod-
ucts:
exo-(ꢀ)-1,8,9,10,11,11-hexachlorotetracyclo[6.2.1.33,6 .02,7]dodecan-4-yl
acetate,
1,8,9,10,11,11-hexachlorohexacyclo[6.2.1.13,6 .02,7.04,9.05,9] dodecane 2, exo-(ꢀ)-1,8,9,10,11,11-
hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-4-yl acetate (ꢀ)-3 and acetate (ꢀ)-89. The chlorinated
birdcage hydrocarbon 2 and chlorinated tetracyclic acetate can be separated by column chromatography,
but chlorinated pentacyclic exo-acetate (ꢀ)-3 and acetate (ꢀ)-8 are very difficult to separate by
this procedure. We have observed that the proportions of derivatives depend on reaction time and
temperature. In 30 min, only the chlorinated birdcage hydrocarbon 2 and acetate pentacyclic exo-(ꢀ)-3
compounds are produced, which are easily separated by column chromatography. By hydrolysis of the
chlorinated pentacyclic exo-acetate with hydrochloric acid in tetrahydrofuran the alcohol (ꢀ)-4 was
obtained.
Scheme 1.
The racemic alcohol (ꢀ)-7 was prepared according to Scheme 2. By epoxidation of isodrin
1 with 3-chloroperoxybenzoic acid in the presence of potassium fluoride,10 the 1,8,9,10,11,11-
hexachlorotetracyclo[6.2.1.33,6.02,7]dodecan-4,5-epoxi-9-eno (endrin) 5 was obtained. The treatment
of endrin with boron trifluoride etherate, following the methodology of Fukanaga,11 produces the

V. E. U. Costa et al. / Tetrahedron: Asymmetry 9 (1998) 2579–2585
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1,8,9,10,11,11-hexachloropentacyclo[6.2.1.13,6 .02,7.05–9]dodecan-4-one (ꢀ)-6, which was reduced by
sodium borohydride giving exclusively the alcohol (ꢀ)-7.
Scheme 2.
Faber et al.12–14 studied the enzymatic resolution of endo-bicyclo[2.2.1]hept-2-yl and endo-
bicyclo[2.2.2]oct-2-yl butyrates and observed that systems with endo-configurations exhibit
enantioselectivity in reactions catalyzed by lipase from Candida rugosa. The R-configuration is
preferentially transesterified.
Lightner et al.15 dechlorinated the tetracyclic alcohol obtained by hydroboration of isodrin, prepared
the half-acid phthalate and resolved it by fractional crystallization of the brucine salt. However, the
enantiomeric excess was only 41.9%. Otherwise, we have observed that the use of enzymes to resolve
dechlorinated polycyclic alcohols does not give good results. The same observation has been found
for alcohols with the exo-configuration. However, enzymatic resolution of 7,7-disubstituted 1,4,5,6-
tetrachlorobicyclo[2.2.1]hept-5-en-2-ols16 using the lipase from Candida rugosa in vinyl acetate was
achieved with complete specificity conversion (∼50%) and high selectivity to the alcohol (ee 98%) and
to the ester (ee >99%).
Encouraged by these findings, we selected the lipase from Candida rugosa for resolving racemic
alcohol (ꢀ)-4 and alcohol (ꢀ)-7 by transesterification with excess vinyl acetate (Scheme 3).
As the enzymatic transesterification of alcohol (ꢀ)-4 did not occur, even with a reaction time of 10
days, the hydrolysis reaction of the respective chlorinated pentacyclic exo-acetate, in aqueous solution of
monobasic potassium orthophosphate (0.01 M) was tested for 7 days, without success. On the other hand,
testing the alcohol (ꢀ)-7 under the same conditions in transesterification (reaction with vinyl acetate, 7
days), a high degree of enantioselectivity of the enzyme was found (E>100)17 and the optically pure
acetate (+)-8 was obtained, as well as the alcohol (−)-7 with high enantiomeric excess. The reaction
yield of enantiomeric conversion to the acetate (+)-8 was 44% and these data are shown in Table 1.
For the determination of the enantiomeric excess of chiral compounds, two techniques are normally
used: (a) chromatography (GC and HPLC) on chiral columns, and (b) NMR spectroscopy. Since
chlorinated compounds have high boiling points, the use of GC on chiral columns with cyclodextrin
is impossible and the use of HPLC on chiral columns with cyclodextrin did not result in satisfactory
separation. Consequently, we have chosen 1H NMR as the analytical method, using chiral chemical shift
reagents. A high resolution of the signals has been achieved for the enantiomeric protons (α-Cl) of both,

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Scheme 3.
Table 1
Results of transesterification catalyzed by lipase from Candida rugosa
alcohol (ꢀ)-7 and acetate (ꢀ)-8 and the CH3 signal of the ester. Sequential addition of 5 mg of tris[3-
(heptafluoropropylhydroxymethylene)-(+)-camphorato] europium(III) (Eu(hfc)3) to CDCl3 solutions of
these compounds (10 mg) in a 5 mm NMR tube, achieved the best results with 20 mg of the chiral
chemical shift reagent. The differences in chemical shifts (∆∆δ) of the enantiomeric hydrogens H10 (α-
Cl) were 0.0074 ppm for the alcohol (ꢀ)-7, 0.041 ppm for the H10 and 0.09 ppm for the CH3 of acetate
(ꢀ)-8 (Fig. 1a, b and c).
3. Experimental section
The melting points were measured on an Electrothermal IA 9100 digital melting point apparatus.
NMR spectra were measured with a Varian VXR-200 spectrometer at a magnetic field of 4.7 T and a
temperature of 22°C. Chemical shifts are expressed as δ (ppm) relative to TMS as an internal standard.
Elemental analyses were recorded on a Perkin–Elmer 2400 CHN elemental analyzer apparatus. Products
were analyzed by GC using capillary columns (25 m×0.2 mm (i.d.)×0.11 µm) packed with SE-30 and a
Varian Model star 3400 CX chromatograph equipped with an FID. Optical rotations were measured with
an Acatec PDA 8200 digital polarimeter with a 1 dm cell at a temperature of 20°C.
Lipase, type VII (from Candida rugosa); tris[3-(heptafluoropropylhydroxymethylene)-(+)-

V. E. U. Costa et al. / Tetrahedron: Asymmetry 9 (1998) 2579–2585
2583
Fig. 1. Spectra: (a) alcohol (ꢀ)-7 and acetate (ꢀ)-8; (b) alcohol (ꢀ)-7 and acetate (ꢀ)-8 in the presence of (+)-Eu(hfc)3; (c)
alcohol (−)-7 and acetate (ꢀ)-8 in the presence (+)-Eu(hfc)3
camphorato] europium(III); chloroform-d1 (99.8 atom% D 98%); 3-chloroperoxybenzoic acid
(50–60%) and isodrin (98%) were purchased from the Aldrich Chemical Co. Vinyl acetate (>99%),
silica gel 60 (230–400 mesh ASTM), sodium borohydride, ethyl acetate (99.5%) and hexane (99%) were
purchased from Merck.
3.1. exo-(ꢀ)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-4-yl acetate (ꢀ)-3
Concentrated sulfuric acid (1 mL) was added to a solution of isodrin (3.0 g, 8.2 mmol) in acetic acid (12
mL) at a temperature of 125°C under magnetic stirring. After 30 minutes, the system was cooled down
and neutralized with a 10% NaHCO3 solution. A precipitate was formed and separated. An additional
extraction with chloroform was undertaken. The extract was combined with the precipitate. This solution
was dried with MgSO4, and after solvent evaporation from the filtrates, a white solid was obtained as a
mixture of two compounds. These products were separated on a silica gel column (hexane:ethyl acetate,
0 to 20%). The obtained compounds were the chlorinated birdcage hydrocarbon 2 (0.8 g, 2.2 mmol),
27% yield, and chlorinated pentacyclic exo-acetate (ꢀ)-3, (1.77 g, 4.1 mmol), 50% yield.
Chlorinated birdcage hydrocarbon 2: mp 290°C (lit.2 287–289°C). 13C NMR (50 MHz, CDCl3): δ
39.8 (CH2), 43.4 (2CH), 54.4 (2CH), 58.4 (2CH), 78.2 (2C), 83.5 (2C), 97.5 (C).
Chlorinated pentacyclic exo-acetate (ꢀ)-3: mp 193–194 (lit.9 194–195). FTIR (film, CHCl3): ν (cm−1
)

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1735 (C_O). 1H NMR (200 MHz, CDCl3): δ 2.1 (s, 3H, methyl), 5.5 (s, 1H, Hα-Cl) and 5.8 (s, 1H, Hα-
O). 13C NMR (50 MHz, CDCl3): δ 21.1 (CH3), 36.5 (CH2), 41.6 (CH), 43.4 (CH), 55.6 (CH), 58.8 (CH),
60.0 (CH), 64.7 (CH), 74.0 (C), 76.4 (CH), 79.6 (C), 84.8 (C), 99.5 (C), 169.8 (C_O).
3.2. exo-(ꢀ)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-4-ol (ꢀ)-4
To a chlorinated pentacyclic exo-acetate (ꢀ)-3 solution (1 g, 2,31 mmol) in THF (50 mL) an aqueous
solution of 15% HCl was added at room temperature under magnetic stirring for 12 hours. The mixture
was neutralized with 10% NaHCO3, extracted with chloroform and this solution was dried with MgSO4.
Evaporation of the solvent gives the respective alcohol (ꢀ)-4 (0.83 g, 2.2 mmol), mp 217 (lit.9 218°C),
95% yield. FTIR (KBr): ν (cm−1) 3471 (C–OH). 1H NMR (200 MHz, CDCl3): δ 5.0 (s, 1H, Hα-O) and
5.4 (s, 1H, Hα-Cl). 13C NMR (50 MHz, CDCl3): δ 36.0 (CH2), 41.2 (CH), 45.3 (CH), 55.6 (CH), 58.9
(CH), 62.3 (CH), 64.6 (CH), 73.7 (CH), 74.3 (C), 79.6 (C), 85.1 (C), 98.6 (C).
3.3. 1,8,9,10,11,11-Hexachlorotetracyclo[6.2.1.33,6 .02,7]dodecan-4,5-epoxi-9-eno (endrin) 5
To an isodrin solution (5.00 g, 13.7 mmol) in CH2Cl2 (15 mL), m-CPBA acid (4.74 g, 27.4 mmol) and
KF (1.6 g, 27.4 mmol) were added. After stirring for 15 hours at r.t., activated KF (1 hour at 120°C/0.1
torr) (1.6 g, 27.4 mmol) was added up to a total 1:2 m-CPBA:KF molar ratio and the reaction mixture
was stirred for 1 hour at r.t. The crude mixture was then filtered off and the solvent was removed by
distillation to give the respective epoxide 5 (4.83 g, 12.69 mmol), mp 246°C dec. (lit.1 245°C dec.), 92%
1
yield. H NMR (200 MHz, CDCl3): δ 3.4 (s, 2H, Hα-O). 13C NMR (50 MHz, CDCl3): δ 29.8 (CH2),
39.2 (2CH), 47.0 (2CH), 54.5 (2CH), 79.5 (2C), 108.7 (C), 132.3 (2C).
3.4. (ꢀ)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6 .02,7.04,5]dodecan-4-one (ꢀ)-6
A solution of endrin 5 (5.27 g, 13.8 mmol) in toluene (20 mL) was heated to reflux (70°C) and then
boron trifluoride etherate (2 mL) in toluene (10 mL) was added. Refluxing was continued for 24 hours.
After cooling a solid precipitated from the reaction mixture. The product was removed by filtration
and cleaned by flash column chromatography (chloroform). Evaporation of the solvent sets free the
pentacyclic ketone (ꢀ)-6 (4.2 g, 11.04 mmol), 80% yield, mp 284–285°C dec. (lit.2 285–286°C dec.).
FTIR (KBr): ν (cm−1) 1750 (C_O). 1H NMR (200 MHz, CDCl3): δ 5.0 (s, 1H, Hα-Cl). 13C NMR (50
MHz, CDCl3): δ 34.3 (CH2), 41.0 (CH), 49.4 (CH), 53.7 (CH), 59.2 (CH), 65.1 (C), 69.0 (CH), 73.3 (C),
79.7 (C), 87.1 (C), 98.8 (C), 205.2 (C_O).
3.5. endo-(ꢀ)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-4-ol (ꢀ)-7
Under an inert gas atmosphere, a mixture of pentacyclic ketone (ꢀ)-6 (4.0 g, 10.47 mmol) and sodium
borohydride (0.70 g, 20.94 mmol) in dry tetrahydrofuran (100 mL) was stirred for 12 hours at r.t. The
resulting anionic complex was decomposed with dilute HCl. The mixture was neutralized with 10%
NaHCO3 and extracted with chloroform. Evaporation of the solvent produces the corresponding alcohol
(ꢀ)-7 (3.16 g, 8.28 mmol), 75% yield, mp 258–260°C dec. (lit.2 258–260°C dec.). FTIR (KBr): ν (cm−1
)
3341 (C–OH). 1H NMR (200 MHz, CDCl3) δ 4.2 (d, J=6.05 Hz, 1H, Hα-OH) and 7.4 (s, 1H, Hα-Cl).
13C NMR (50 MHz, CDCl3): δ 38.0 (CH2), 42.9 (CH), 44.1 (CH), 56.0 (CH), 56.3 (CH), 58.5 (CH),
64.6 (CH), 72.7 (C), 79.8 (CH), 81.1 (C), 83.3 (C), 99.3 (C).

V. E. U. Costa et al. / Tetrahedron: Asymmetry 9 (1998) 2579–2585
2585
3.6. endo-(+)-1,8,9,10,11,11-Hexachloropentacyclo[6.2.1.13,6 .02,7.05,9]dodecan-4-yl acetate (+)-8
Lipase (50% w/w of substrate) was added to a solution of pentacyclic alcohol (ꢀ)-7 (0.38 g, 1 mmol) in
vinyl acetate (10 mL) and the suspension was shaken at 250 rpm at 25°C. When the appropriate degree of
conversion was achieved (7 days), the enzyme was filtered and the excess vinyl acetate was evaporated.
The products were separated by flash column chromatography (hexane:ethyl acetate=4:1), giving the
alcohol (−)-7 and acetate (+)-8 (0.19 g, 0.44 mmol), 44% yield.
Alcohol (−)-7: mp 258–260°C dec., [α]D −0.86 (c 0.6, ethyl acetate). FTIR (KBr): ν (cm−1) 3341
20
1
(C–OH). H NMR (200 MHz, CDCl3) δ 4.2 (d, J=6.05 Hz, 1H, Hα-OH) and 7.4 (s, 1H, Hα-Cl). 13C
NMR (50 MHz, CDCl3): δ 38.0 (CH2), 42.9 (CH), 44.1 (CH), 56.0 (CH), 56.3 (CH), 58.5 (CH), 64.6
(CH), 72.7 (C), 79.8 (CH), 81.1 (C), 83.3 (C), 99.3 (C). Anal. calcd: C, 37.60; H, 2.61. Found: C, 37.58;
H, 2.59.
Acetate (+)-8: mp 153–154°C, [α]D +0.75 (c 2, ethyl acetate). FTIR (KBr): ν (cm−1) 1731 (C_O).
20
1H NMR (200 MHz, CDCl3) δ 2.19 (s, 3H, CH3), 4.92 (d, J=6.05 Hz, 1H, Hα-OAc) and 6.57 (s, 1H,
Hα-Cl). 13C NMR (50 MHz, CDCl3): δ 21.0 (CH3), 37.4 (CH2), 42.5 (CH), 43.1 (CH), 54.8 (CH), 56.1
(CH), 58.6 (CH), 64.8 (CH), 72.4 (C), 79.5 (CH), 80.6 (C), 83.4 (C), 99.0 (C), 169.4 (C_O). Anal. calcd:
C, 39.53; H, 2.82. Found: C, 39.77; H, 2.79.
Acknowledgements
The authors thank CNPq, CAPES and FAPERGS for financial support.
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