Carboxylester-Lipase-Mediated Reactions - ACS Symposium Series

Chapter 4

Carboxylester-Lipase-Mediated Reactions A Versatile Route to Chiral Molecules Detlef Lutz, Manfred Huffer, Doris Gerlach, and Peter Schreier

Downloaded by UNIV OF ARIZONA on October 16, 2015 | http://pubs.acs.org Publication Date: May 13, 1992 | doi: 10.1021/bk-1992-0490.ch004

Lehrstuhl Für Lebensmittelchemie, Universität Würzburg, Am Hubland, D-8700 Würzburg, Germany

Two classes of naturally occurring flavor compounds were prepared in optically enriched forms by means of lipase catalyzed transformations in organic solvents, i.e. γ- and δlactones as well as secondary alcohols. The lactones were obtained via intramolecular transesterification reactions, while the chi ral secondary alcohols were prepared via kine tic resolution using esterification with an achiral acid. For the latter compounds a "mo del" was developed describing reactivity and selectivity of the enzymatic catalysis, thus providing a powerful tool to establish future synthetic strategies.

C h i r a l s e c o n d a r y a l c o h o l s , e s t e r s and l a c t o n e s a r e w i d e s p r e a d compounds o f t e n f o u n d i n f o o d f l a v o r s (1). They comprise saturated, u n s a t u r a t e d and a r o m a t i c alcohols, t h e i r f a t t y a c i d e s t e r s , and γ - a n d δ - l a c t o n e s , o c c u r r i n g in d i s t i n c t enantiomeric r a t i o s i n nature (2-5). Their production i n preparative scale is of interest due to d i f f e r e n t reasons: (i) I f these substances are obtained i n o p t i c a l l y e n r i c h e d or pure forms, a n a l y t i c a l p r o p e r t i e s may be e v a l u a t e d , i . e . e n a b l i n g us t o e s t a b l i s h t h e i r a b s o l u t e c o n f i g u r a t i o n s or to determine the o r d e r of e l u t i o n i n c h i r a l chromatography, ( i i ) A f t e r the s e p a r a t i o n of the enantiomers i t i s a l s o p o s s i b l e to c h a r a c t e r i z e t h e i r se lective sensory p r o p e r t i e s (6). (iii) Finally, "biosynt h e t i c " p r e p a r a t i o n s o f i n d u s t r i a l l y a t t r a c t i v e compounds with n a t u r a l enantiomeric r a t i o s are v e r s a t i l e ingredients i n food f l a v o r i n g ( 7 , 8 ) .

0097-6156/92/0490-0032$06.00/0 © 1992 American Chemical Society

In Flavor Precursors; Teranishi, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1992.

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33

Carboxylester-Lipase-Mediated Reactions


What a r e t h e t e c h n i q u e s t o g e t a t a r g e t m o l e c u l e h i g h l y o p t i c a l l y e n r i c h e d ? B e s i d e s c h e m i c a l methods w h i c h a r e often sophisticated (9), and c h r o m a t o g r a p h i c means (10), w h i c h a r e c o n n e c t e d w i t h h i g h c o s t s , t h e r e i s a t h i r d way b a s e d on b i o t r a n s f o r m a t i o n . S i n c e K l i b a n o v ' s f i r s t r e p o r t on t h e c a t a l y t i c a c t i v i t y o f enzymes i n o r g a n i c s o l v e n t s ( 1 1 ) , e n z y m a t i c r e a c t i o n s h a v e become more and more i m p o r t a n t i n the s y n t h e s i s of f i n e chemicals (12). How i s i t p o s s i b l e t o o b t a i n o p t i c a l l y e n r i c h e d a l c o h o l s , e s t e r s and l a c t o n e s by means o f enzyme m e d i a t e d b i o c o n v e r s i o n s ? T h e r e a r e t h r e e g e n e r a l pathways t o u s e a c a r b o x y l e s t e r l i p a s e system t o achieve t h i s aim, i . e . (i) hydrolysis, ( i i ) e s t e r i f i c a t i o n , and ( i i i ) transesterifi c a t i o n . From t h e methods shown i n F i g u r e 1 we h a v e u s e d b o t h t h e e s t e r i f i c a t i o n and t r a n s e s t e r i f i c a t i o n i n o r g a n i c solvents. I n t h e f o l l o w i n g , t h e p r o d u c t i o n o f γ - and δ lactones a s w e l l a s s e c o n d a r y a l c o h o l s by c a r b o x y l e s t e r lipase is described.

Enzyme

Selection

F i r s t o f a l l , a s c r e e n i n g was p e r f o r m e d t o s e l e c t t h e most a p p r o p r i a t e c a r b o x y l e s t e r l i p a s e . A number o f p r e p a r a t i o n s are c o m m e r c i a l l y a v a i l a b l e i n c l u d i n g c a t a l y s t s from m i c r o b i a l , p l a n t , and a n i m a l s o u r c e s . They were c h e c k e d i n an esterification system c o n s i s t i n g of (R,S)-2-octanol and d o d e c a n o i c a c i d i n n - h e x a n e ( 1 3 ) . The r e s u l t s a r e s u m m a r i z e d i n F i g u r e 2. From t h e s i x p r e p a r a t i o n s s t u d i e d o n l y two p r o v i d e d s a t i s f y i n g f i n d i n g s . The enzyme f r o m Candida cylindracea showed a c o n v e r s i o n r a t e o f a p p r o x i m a t e l y 70 %, b u t e x h i b i t e d v e r y low s e l e c t i v i t y . As t o t h e selecti v i t y - the p r i o r c r i t é r i u m f o r our s e l e c t i o n - the e x t r a c t f r o m p i g p a n c r e a s (PPE) was s u p e r i o r t o a l l o t h e r t e c h n i c a l enzymes (ee = 85 %). T h i s p r e p a r a t i o n , w h i c h showed a c o n v e r s i o n r a t e o f a p p r o x i m a t e l y 20 %, was s e l e c t e d f o r further studies. I t s h o u l d be n o t e d t h a t t h e (R)-enant i o m e r was p r e f e r a b l y e s t e r i f i e d by a l l t h e enzyme p r e p a r a t i o n s under study. Production of O p t i c a l l y Enriched

Lactones

The r e a c t i o n pathway l e a d i n g t o o p t i c a l l y e n r i c h e d l a c t o n e s i s o u t l i n e d i n F i g u r e 3 . The i n t e r e s t i n g compounds, i.e. γ - and δ - l a c t o n e s w i t h c h a i n l e n g t h s f r o m C t o C were c h e m i c a l l y c o n v e r t e d i n t o t h e c o r r e s p o n d i n g h y d r o x y propylesters (14) t h a t were s u b s e q u e n t l y t r a n s f o r m e d i n organic solvent via i n t r a m o l e c u l a r transesterification u s i n g P P E . C h i r a l a n a l y s i s o f t h e h y d r o x y e s t e r s was p e r f o r m e d by t h e i r d e r i v a t i z a t i o n i n t o d i a s t e r e o m e r s using chiral agents, such as (R)-(+)-a-methoxy-a-trifluoromethylphenylacetylchloride ((R)-MTPA-Cl), (R)-(+)-a-phen y l e t h y l i s o c y a n a t e ( ( R ) - P E I C ) , and ( S ) - O - a c e t y l l a c t i c a c i d ((S)-ALA-C1) (15), a n d by s u b s e q u e n t g a s c h r o m a t o g r a p h i c 5


1

3

FLAVOR PRECURSORS

Hydrolysis : R-COOR' + H 0

1

R-COOH + R -OH

2


Esterification: R-COOH + R'-OH

R-COOR + H 0 1

2

Transesterification: 1

f

R-COOR ' + R -OH

F i g u r e 1.

100

Scheme o f

•

1

R-COOR' + R *-OH

lipase catalyzed reactions.

% ee (R)

% conversion

Aspergillus niger

Rhizopue arrhizue

Candida cylindracea

I % conversion

Mucor miehei

H

PiQ pancreas

100

Wheat germ

% ee (R)

F i g u r e 2. S c r e e n i n g o f l i p a s e s by e s t e r i f i c a t i o n o f ( R , S ) - 2 - o c t a n o l and d o d e c a n o i c a c i d i n hexane a t 4 0 ° C (24 h) (13).


4. LUTZ ET A L


35

s e p a r a t i o n (HRGC). A n a l y s i s o f t h e o b t a i n e d l a c t o n e s was c a r r i e d o u t w i t h o u t d e r i v a t i z a t i o n by means o f h i g h p e r formance liquid chromatography (HPLC) using ChiraSpher (Merck) as c h i r a l s t a t i o n a r y p h a s e (10). The r e s u l t s o b t a i n e d by t h e e n z y m a t i c t r a n s f o r m a t i o n o f 4-hydroxyesters are i l l u s t r a t e d i n Figure 4 f o r the C and Cn-homologues. In a l l cases the r e s u l t i n g lactones were ( S ) - c o n f i g u r a t e d , w h e r e a s t h e r e m a i n i n g h y d r o x y e s t e r s had ( R ) - c o n f i g u r a t i o n ( t h e y c o u l d be r e a r r a n g e d c h e m i c a l l y to y i e l d the corresponding (R)-lactones). Downloaded by UNIV OF ARIZONA on October 16, 2015 | http://pubs.acs.org Publication Date: May 13, 1992 | doi: 10.1021/bk-1992-0490.ch004

5

The following conclusions c a n be drawn f o r t h e PPE mediated p r o d u c t i o n of γ - l a c t o n e s : ( i ) h i g h o p t i c a l y i e l d s (ee = > 80 %) were o b t a i n e d f o r e i t h e r l a c t o n e o r h y d r o x y e s t e r ; ( i i ) m o d e r a t e r e a c t i o n r a t e s ( 1 0 - 4 0 h) were o b s e r ved, t h e l o n g - c h a i n homologues r e a c t i n g f a s t e r t h a n t h e s h o r t - c h a i n compounds; ( i i i ) a f t e r h a v i n g r e a c h e d a m a x i mum v a l u e , t h e ee v a l u e s o f t h e l a c t o n e s d e c r e a s e d s l i g h t l y due t o c h e m i c a l r e l a c t o n i z a t i o n o f t h e e s t e r s . The r e s u l t s o b t a i n e d f r o m t h e PPE c a t a l y z e d c o n v e r s i o n s of 5-hydroxyesters are o u t l i n e d i n Figure 5 f o r the C n homologue. In comparison to the 4-hydroxyesters the r e a c t i o n r a t e s determined f o r the 5-hydroxyesters were f i v e t o s i x t i m e s l o w e r . I n a d d i t i o n , low enantioselec t i v i t y (ee < 20 %) was o b s e r v e d . T h e r e f o r e , t h e p r e p a r a t i o n o f o p t i c a l l y e n r i c h e d δ - l a c t o n e s via t r a n s e s t e r i f i c a t i o n of 5-hydroxyesters i s s t r o n g l y l i m i t e d by t h e sub s t r a t e s p e c i f i c i t y of PPE.

Production of Optically Enriched Alcohols By r e a c t i o n o f a r a c e m i c a l c o h o l w i t h an a c h i r a l fatty a c i d under l i p a s e c a t a l y s i s the a l c o h o l enantiomers are converted at d i f f e r e n t reaction rates, r e s u l t i n g i n op t i c a l e n r i c h m e n t o f t h e produced e s t e r and t h e r e m a i n i n g a l c o h o l ( k i n e t i c r e s o l u t i o n , c f . F i g . 6) (16). In o u r s t u d y t o p r o d u c e r e l e v a n t f l a v o r a l c o h o l s and e s t e r s by k i n e t i c r e s o l u t i o n , we were s u c c e s s f u l t o e s t a b l i s h a "model" d e s c r i b i n g r e a c t i v i t y and s e l e c t i v i t y of t h e PPE c a t a l y z e d e s t e r i f i c a t i o n i n o r g a n i c media (17). S i m i l a r d e s c r i p t i o n s o f enzymic a c t i v i t i e s have been p r o v i d e d by Ohno e t a l . (18) u s i n g p i g l i v e r e s t e r a s e ( P L E ) and, r e c e n t l y , by E h r l e r and S e e b a c h (19) i n t h e i r r e p o r t on t h e s e l e c t i v i t y o f PPE i n w a t e r s y s t e m s . F i r s t o f a l l , s t a n d a r d e x p e r i m e n t a l c o n d i t i o n s were d e v e l o p e d t o e f f i c i e n t l y e s t e r i f y c h i r a l ( R , S ) - a l c o h o l s . The f o l l o w i n g r e a c t i o n c o n d i t i o n s were c h o s e n f o r a l l t r a n s f o r m a t i o n s : To a s o l u t i o n o f 2 . 5 mmol o f e a c h r a c e m i c a l c o h o l and d o d e c a n o i c a c i d i n 25 ml n - h e x a n e 1 g PPE was a d d e d and t h e m i x t u r e s t i r r e d a t 7 0 ° C f o r 24 h o u r s . A f t e r



FLAVOR

PRECURSORS

4-hydroxyester PPE

S-lactone

R-4-hydroxyester

F i g u r e 3. R e a c t i o n pathway l e a d i n g t o o p t i c a l l y e n r i c h e d γ - l a c t o n e s . PPE = p o r c i n e p a n c r e a s e x t r a c t (14,15).

%

0

10

20

30 40 50 reaction time (h)

60

70

80

-β- % conversion (C-5) —·— %ee R-ester (C-5) - * - %ee S-lacton (C-5) % conversion (C-11) - * ~ %ee R-ester (C-11)

%ee S-lacton (C-11)

F i g u r e 4. PPE c a t a l y z e d t r a n s e s t e r i f i c a t i o n o f hydroxyesters i n d i e t h y l ether at 20°C (15).


4-

4. LUTZ ET AL.


37

% 100-Γ" 80-


60402004 0

2

4 6 reaction time (d)

— ^ % conversion (C-11)

—

h _

8

10

%ee R-ester (C-11)

%ee S-lacton (C-11)

F i g u r e 5. PPE c a t a l y z e d t r a n s e s t e r i f i c a t i o n o f hydroxyesters i n d i e t h y l ether at 20°C (15).

F i g u r e 6. Lipase catalyzed e s t e r i f i c a t i o n (R,S)-alcohol (16).

of


5-

an

38

FLAVOR

PRECURSORS


cooling, the enzyme was f i l t e r e d off and t h e reaction m i x t u r e worked up f o r t h e a n a l y s i s o f p r o d u c t s and e d u c t s . Esters were r e d u c t i v e l y cleaved yielding the original a l c o h o l s w h i c h c o u l d be e a s i l y a n a l y z e d u s i n g t h e same d e r i v a t i z a t i o n a g e n t s as mentioned p r e v i o u s l y f o r t h e h y d roxyesters o r u s i n g t h e c h i r a l L i p o d e x C column w i t h o u t any d e r i v a t i z a t i o n (17). C o n f i g u r a t i o n s were determined with the help of polarimetry, i n case l i t e r a t u r e data existed, o r by NMR e x p e r i m e n t s , if t h o s e were l a c k i n g (20). F o r more d e t a i l e d a n a l y t i c a l i n f o r m a t i o n see refe rence (17).

a.

2-Alkanols

and

Ε - 3 - Al ken-2 - ο 1 s

The r e s u l t s o b t a i n e d i n t h e homologous s e r i e s o f 2 - a l k a n o l s and E - 3 - a l k e n - 2 - o l s a r e o u t l i n e d i n T a b l e I . The c o n v e r s i o n r a t e s f o r the s a t u r a t e d s u b s t r a t e s ranged from 20-30 %, w h e r e a s t h o s e o f t h e u n s a t u r a t e d a n a l o g s were a b o u t 40 %. I t seemed t h a t t h e a l k e n o l s were a c t i v a t e d by t h e i n f l u e n c e o f t h e d o u b l e bond. As t o t h e selectivity, t h e r e was no s i g n i f i c a n t d i f f e r e n c e between t h e two c l a s ses. In a l l c a s e s , the p r e f e r r e d e s t e r i f i e d a l c o h o l was ( R ) - c o n f i g u r a t e d w i t h c o n s i d e r a b l y h i g h ee v a l u e s (about 90 %). Table I. PPE c a t a l y z e d E-3-alken-2-ols Substrate

% Ester

esterification

ee E s t e r ee

of

2-alkanols

Alcohol Preferred

2-Pentanol 2-Hexanol 2-Heptanol 2-Octanol 2-Nonanol

21.2 30.1 27.0 31.3 19.3

87.4 95.9 92.3 93.1 92.7

26.8 33.8 33.9 41.4 25.1

R R R R R

E-3-Penten-2-ol E-3-Hepten-2-ol E-3-Octen-2-ol

43.0 40.3 42.6

79.1 86.1 93.8

77.6 97.0 97.7

R R R

b.

1 - Aiken-3

and

Antipode

-ols

Conversion r a t e s f o r the l - a l k e n - 3 - o l s r a n g e d f r o m 15-30 %. The r e s u l t s o b t a i n e d i n t h i s s e r i e s c o n c e r n i n g s e l e c tivity are i l l u s t r a t e d i n F i g u r e 7. As shown f r o m t h e graph, the c o n f i g u r a t i o n of the p r e f e r a b l y converted enant i o m e r s c h a n g e d between t h e C and t h e C a l k e n o l . S u c h a r e v e r s a l o f c o n f i g u r a t i o n has b e e n p r e v i o u s l y o b s e r v e d i n PLE c a t a l y z e d h y d r o l y s e s (21). For better understanding t h i s phenomenon, t h e s t r u c t u r e s o f t h e p r e f e r r e d e n a n t i o mers a r e o u t l i n e d a s f o l l o w s : 5

6


4. LUTZ ET AJL In t h i s


p r o j e c t i o n , the

shorter,

less bulky side

39

chains


H

a r e i n t h e t o p p o s i t i o n , whereas t h e l o n g e r ( l a r g e r ) s i d e c h a i n s a r e a t t h e b o t t o m . I n t h e homologous s e r i e s the r e v e r s a l o f c o n f i g u r a t i o n o c c u r r e d when b o t h s i d e c h a i n s were s t e r i c a l l y e q u a l , i . e . i n t h e c a s e o f l - p e n t e n - 3 - o l . A t t h e same t i m e t h e a b s o l u t e ee v a l u e r e a c h e d a minimum. The p r e l i m i n a r y "model" o f t h e r e a c t i v e e n a n t i o m e r s as described above should be approved i n the following studies. C·

1-Aiken-4-ols

F i g u r e 8 shows t h e r e s u l t s o f t h e c o n v e r s i o n s o b t a i n e d i n the s e r i e s of l - a l k e n - 4 - o l s . Again a r e v e r s a l of the conf i g u r a t i o n was o b s e r v e d when t h e s i d e c h a i n s became s t e r i c a l l y e q u a l , i . e . between t h e C and t h e C homologue. The r e a c t i o n r a t e s d e c r e a s e d f r o m 36 % ( l - p e n t e n - 4 - o l ) to 20 % ( l - o c t e n - 4 - o l ) , an e f f e c t t h a t became e v e n more d r a s t i c i n the s e r i e s of E - 2 - a l k e n - 4 - o l s . 6

d.

7

E-2-Alken-4-ols

In t h i s s e r i e s of experiments, a decrease of e s t e r form a t i o n r a t e w i t h i n c r e a s i n g c h a i n l e n g t h was observed (Table II). The r e d u c e d a c c e s s i b i l i t y of the reactive hydroxy group, which i s s i t u a t e d i n the middle of the m o l e c u l e s i n t h e c a s e o f t h e h i g h e r h o m o l o g u e s , may be responsible for this effect. In analogy t o the l - a l k e n - 3 o l s and l - a l k e n - 4 - o l s a r e v e r s a l o f c o n f i g u r a t i o n o f t h e p r e f e r a b l y c o n v e r t e d e n a n t i o m e r s was f o u n d , a g a i n a t t h e p o s i t i o n where b o t h s i d e c h a i n s were s t e r i c a l l y e q u a l , i . e . between C and C . 7

e·

Isomeric

8

Hexenols

Not o n l y i n homologous s e r i e s , but a l s o i n the case of i s o m e r i c compounds t h e phenomenon o f r e v e r s a l o f c o n f i g u r a t i o n was o b s e r v e d a s i l l u s t r a t e d i n F i g u r e 9 . Due t o the p r e v i o u s l y mentioned r e a s o n , i . e . the a c c e s s i b i l i t y of t h e r e a c t i v e g r o u p , l - h e x e n - 5 - o l was f o u n d t o be t h e most r e a c t i v e r a c e m a t e i n t h i s s e r i e s ( c o n v e r s i o n r a t e , 43 %).


40

FLAVOR PRECURSORS


% ee of the reactive enantiomers

4

5

6

7

8

9

Number of C-atoms

Figure 7. PPE catalyzed esterification of homologous l-alken-3-ols. (Reproduced with permission from reference 17. Copyright 1990 Pergamon.)

% ee of the reactive enantiomers

5

6

7

8

Number of C-atoms

Figure 8. PPE catalyzed esterification of homologous l-alken-4-ols. (Reproduced with permission from reference 17. Copyright 1990 Pergamon.)



4. LUTZ ET AL.


% ee of the reactive enantiomers 60 ,

40

20

0

20

h

40

l.

60

Y

80

1

1

1-hexen-3-ol

' 1-hexen-4-ol

1

1-hexen-5-ol

Figure 9. PPE catalyzed esterification of isomeric hexenols. (Reproduced with permission from reference 17. Copyright 1990 Pergamon.)


41

42

FLAVOR

Table

II.

PPE c a t a l y z e d

Substrate


f.

Ζ/Ε - Isomer

43.0 33.7 12.0 5.7

ic

of

E-2-alken-4-ols

ee E s t e r ee A l c o h o l P r e f e r r e d

% Ester

E-2-Penten-4-ol E-2-Hexen-4-ol E-2-Hepten-4-ol E-2-Octen-4-ol

esterification

PRECURSORS

R R R S

77.6 60.8 11.1 1.1

79.1 90.0 71.9 80.0

Antipode

Alkenols

The e s t e r f o r m a t i o n r a t e s o f Z - i s o m e r s were d e t e r m i n e d t o be c o n s i d e r a b l y l o w e r t h a n t h o s e o f t h e a n a l o g o u s E - i s o mers ( T a b l e I I I ) . O b v i o u s l y , the r e a c t i v e center i s p r o t e c t e d by t h e p a r t i c u l a r g e o m e t r y o f t h e a l l y l i c Z - d o u b l e b o n d . I n a l l c a s e s , t h e ( R ) - e n a n t i o m e r was t h e p r e f e r r e d substrate. However, t h e e s t e r i f i c a t i o n of E-isoraers ex h i b i t e d much h i g h e r s e l e c t i v i t y w h i c h i s a l s o an e f f e c t o f s u b s t r a t e and enzyme g e o m e t r y .

T a b l e I I I . PPE c a t a l y z e d gurated alkenols Substrate

% Ester

esterification

of

Z - and

E-confi-

ee E s t e r ee A l c o h o l P r e f e r r e d

Antipode

Z-3-Hepten-2-ol E-3-Hepten-2-ol

6.9 40.3

53.3 86.1

1.2 97.0

R R

Z-3-Octen-2-ol E-3-Octen-2-ol

14.3 42.3

40.6 93.8

9.1 97.7

R R

g·

Aromat

ic

Alcohols

I n t h i s c l a s s o f compounds n e a r l y c o n s t a n t and h i g h ee v a l u e s ( a b o u t 90 %, c f . T a b l e I V ) were f o u n d . O b v i o u s l y , t h e a r o m a t i c r i n g s i n t h e s u b s t r a t e s a r e t h e main f a c t o r s determining the selectivity. In the above illustrated s t e r e o c h e m i c a l p r o j e c t i o n ( c f . b . ) t h e l a r g e benzene r i n g w o u l d be a t t h e b o t t o m p o s i t i o n , i.e. a l s o the aromatic s u b s t r a t e s a r e i n a c c o r d a n c e w i t h the p o s t u l a t e d "model". The c o n v e r s i o n r a t e s m e a s u r e d were d e p e n d e n t on t h e p o s i t i o n o f t h e h y d r o x y g r o u p ( C o r C ) and r a n g e d f r o m 20 t o 40 %. 2

3


4. LUTZ ET AK Table IV. cohols


Substrate

Carboxylester-Lipase-Mediated Reactions PPE c a t a l y z e d

% E s t e r ee

1-Phenyl1-ethanol l-Phenyl-2propen-l-ol 1-Phenyl1-propanol 4-Phenyl-E-3 buten-2-ol 4-Phenyl2-butanol

esterification

E s t e r ee

of

43

aromatic

al-

Alcohol Preferred Antipode

33.0

92.7

52.7

R

19.3

85.7

30.0

R

18.0

86.9

19.6

R

43.8

88.5

73.7

R

28.9

83.3

44.4

R

Conclusions As a g e n e r a l r e s u l t i t c a n be s t r e s s e d t h a t t h e esterified enantiomers e x h i b i t the following tions :

preferably configura-

OH

Ri i s the s h o r t e r , side chain.

less

bulky,

R

2

the

longer

or

larger

The k i n e t i c r e s o l u t i o n o f a r a c e m a t e i s a two s u b s t r a t e r e a c t i o n . I f we l o o k a t t h e r e a c t i v i t y o f a s i n g l e e n a n t i o m e r , i t c a n be r u l e d o u t t h a t a s h o r t R w i l l r e s u l t i n high r e a c t i v i t y ; i n the optimal case R i s a methyl group. T h e r e f o r e , t h e most r e a c t i v e compounds a r e . . . 2 - o l s . C o n versely, large R l i k e a b e n z e n e r i n g , w i l l l e a d t o low r e a c t i v i t y . T h i s may be i l l u s t r a t e d by two e x a m p l e s : x

x

lf

OH

OH

A ^

A C H 6

U

(R)-E-3-octene-2-ol ( 5.2 % conversion )

P

h

. c

2

H ?

OT

>

(S)-4-phenyl-E-3-butene-2-ol ( 98.9 % conversion )


44

FLAVOR PRECURSORS


A racemate i s transformed w i t h h i g h e n a n t i o s e l e c t i v i t y , if t h e two e n a n t i o m e r s d i f f e r s t r o n g l y i n t h e i r r e a c t i v i t i e s , i.e. t h e two s i d e c h a i n s h a v e t o be f a i r l y d i f f e r e n t in t h e i r s t e r i c a l d i m e n s i o n s . A good example f o r t h i s s t a t e ment i s ( R , S ) - 2 - o c t a n o l w h i c h i s e s t e r i f i e d w i t h an e n a n t i o s e l e c t i v i t y o f 93 % e e .

However, some i n c o n s i s t e n c i e s were f o u n d , i.e. the r e l a t i v e l y h i g h ee v a l u e s d e t e r m i n e d f o r l-hepten-3-ol. But i t s h o u l d be c o n s i d e r e d t h a t t h e s t u d i e s were c a r r i e d out w i t h a crude p r o t e i n e x t r a c t . In o r d e r t o c h a r a c t e r i z e t h e a c t i v e e n z y m e ( s ) a c c o r d i n g t o i t s ( t h e i r ) k i n e t i c and structural features, purification is needed. Actually, there a r e some h i n t s that the common t r i a c y l g l y c e r i d e l i p a s e of p i g pancreas i s not r e s p o n s i b l e f o r the catal y s i s i n o r g a n i c m e d i a ( 2 2 , 2 3 ) . Work t o c h a r a c t e r i z e t h e e f f e c t i v e enzyme i s i n p r o g r e s s i n o u r l a b o r a t o r y .

Acknowledgement.

The f i n a n c i a l s u p p o r t o f t h i s work by t h e Deutsche Forschungsgemeinschaft, Bonn, i s g r a t e f u l l y a c k n o w l e d g e d . The a u t h o r s t h a n k A . G u l d n e r and R . Thums f o r their helpful collaboration.

LITERATURE CITED 1. Bauer, K.; Garbe, D.; Surburg, H. Common Fragrance and Flavor Materials, 2nd. ed., VCH Verlagsgesellschaft, Weinheim, 1990. 2. Engel, K.H. In: Bioflavour '87, Schreier, P., ed.; W. de Gruyter: Berlin, New York, 1988, pp. 89-95. 3. Fröhlich, O.; Huffer, M.; Schreier, P. Z. Naturforsch. 1989, 44c, 555. 4. Mosandl, A. Food Rev. Intern. 1988, 4, 1. 5. Bernreuther, A.; Christoph, N.; Schreier, P. J. Chromatogr. 1989, 481, 363. 6. Mosandl, Α.; Heusinger, G.; Gessner, M. J. Agric. Food Chem. 1986, 34, 119. 7. Drawert, F. In: Bioflavour '87, Schreier, P., ed., W. de Gruyter: Berlin, New York, 1988, pp. 3-32. 8. Schreier, P. Food Rev. Intern. 1989, 5, 289. 9. Helmchen, G.; Ihrig, K.; Schindler, H. Tetrahedron Lett. 1987, 28, 183. 10. Huffer, M.; Schreier, P. J. Chromatogr. 1989, 469, 137.



4. LUTZ ET AK


45

11. Cambou, B.; Klibanov, A.M. J. Am. Chem. Soc. 1984, 106, 2687. 12. Crout, D.H.G.; Christen, M. In: Modern Synthetic Methods 1989, Scheffold, C.R., ed., Springer: Berlin, Heidelberg, New York, pp. 1-114. 13. Gerlach, D. Doctoral Thesis, University of Würzburg, 1988. 14. Mosandl, Α.; Günther, C. J. Agric. Food Chem. 1989, 37, 413. 15. Huffer, M. Doctoral THesis, University of Würzburg, 1990. 16. Chen, C.S.; Fujimoto, Y.; Girdaukas, G.; Sih, C . J . J. Am. Chem. Soc. 1982, 104, 7294. 17. Lutz, D.; Güldner, Α.; Thums, R.; Schreier, P. Tetrahedron Asymmetry 1990, 1, 783. 18. Ohno, M.; Kobayashi, S.; Adachi, K. In: Enzymes as Catalysts in Organic Synthesis, Schneider, M.P., ed.; Reidel: Dordrecht, 1986, pp. 123-142. 19. Ehrler, J.; Seebach, D. Liebigs Ann. Chem. 1990, 379. 20. Helmchen, G.; Schmierer, R. Angew. Chem. 1976, 13, 770. 21. Björkling, F . ; Boutelje, J.; Gatenbeck, S.; Hult, Κ.; Norin, T . ; Szmulik, P. Tetrahedron 1985, 41, 1347. 22. Tombo, G.M.R.; Schaer, H.P.; Fernandez I Busquets, X.; Ghisalba, O. Tetrahedron Lett. 1986, 27, 5707. 23. Hemmerle, H.; Gais, H.J. Tetrahedron Lett. 1988, 28, 3471. RECEIVED December 18, 1991


Carboxylester-Lipase-Mediated Reactions - ACS Symposium Series

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