Metode de obtinere a compusilor enantiomeric puri

Chiral pools Racemates Prochiral substrates

resolution
synthesis asymmetric synthesis
kinetic chromatography
crystallization (bio)catalysis
enzymatic chemical

Enantiomerically pure compunds

 Rezolutia cinetica enzimatica(EKR)
-este realizarea a unei rezolutii partiale sau complete, bazate pe diferentele intre vitezele
de reactie a enatiomerilor dintr-un racemat in interactiunea cu un biocatalizator;
-rezolutia cinetica enzimatica se bazeaza pe preferinta unei enzime pentru unul dintre
enantiomerii substratului (enantiomer selectivitate)
Avantaje: -exces enantiomeric foarte mare;
-amestecul recemic este mai ieftin;
-produsii se pot separa foaret usor;
Dezavantaj:-cel mai important dezavantaj al acestei tehnici este ca maximul
randamentului teoretic este de 50% datorita consumarii doar al unui enantiomer;
- atunci cand catalizatorul este enantioselectiv, cei doi enantiomeri sunt transformati in
produsii corespunzatori cu viteze de reactie diferite.
-Raportul vitezelor de reactie (vA and vB) se numeste raportul enantiomeric
(enantiomeric ratio) (E), si reprezinta enantioselectivitatea enzimei pentru cei doi
enantiomeri.
- Deoarece viteza de reactie este caracterizata de Vmax si Km, E poate fi exprimat cu
urmatoarea ecuatie(1):
 100 100
ees ees
% eep %
eep E=5 ees - Cu ct valoarea lui E este mai mare, crete puritatea
eep
E=20 optic a produsului la aceeai valoare a conversiei.
- E<15 sunt inacceptabile practic,
50 50 -sistemele cu 15 < E < 30 sunt considerate bune
-E peste 30 sunt considerate excelente.
-Practic evaluarea lui E devine dificil la valori ale lui
E>200, deoarece schimbri mici n valorile ees sau eep
duc la modificri mari ale lui E.
0 0
0 50 100 0 50 100
c o n v e rs ia % c o n v e rs ia %
A. B.
Dependena excesului enantiomeric al produsului i al substratului
de conversie n cazul unui proces ireversibil

k1 k3
A EA E+P
ln[1 c(1 eep)] ln[1 c(1 ees)]
k2 E sau E
ln[1 c(1 eep)] ln[1 c(1 ees)]
k4 k6
B EB E+Q
k5 [A] [B] [ P ] [ Q] [B] [A]
c 1 , eep iar ees
[A 0 ] [B0 ] [ P ] [ Q] [ B] [A]
Modelul cinetic al unui proces ireversibil

C= ees/(ees+eep)

ees, eep- excees enantiomeric

C- conversia
 Kinetic resolution of a racemic mixture

OAc OH OAc
lipase
+
buffer
R,S-acetate (S)-alcohol (R)-acetate
racemate
50% y 50%y

Chiral recognition by lipases

The fast reacting enantiomer (a) and the slow reacting one (b) in the active side model for lipases derived from Kazlauskas rule

- the fast-reacting enantiomer binds to the active side in the manner shown
in Figure a; however, when the other enantiomer reacts with the lipase, it is
forced to accommodate its large substituent into the smallest pocket (Fig. b).
 Lipaze
Lipazele: -sunt serinproteaze care pot cataliza hidroliza i sinteza de
acilgliceroli. Aceste reacii decurg adesea cu o regioselectivitate i o
enantioselectivitate mare, astfel lipazele au devenit un foarte
important biocatalizator folosit n chimia organic.
Surse de lipaza:
-surse animale
-surse vegetale
-surse microbiene
-bacteriene, drojdii(yeast), fungi;

CRL1

http://www.au-kbc.org/beta/bioproj2/gall.html
Microorganisms cited in the recent literature as
potential lipase producers
- Lipases are the most versatile of enzymes.
- They are used in a number of bioconversions and their applications can be found in industries like
pharmaceutical, dairy, detergent, cosmetic, oleochemical and others.

O O
A R' + H2O H + R'OH
R O R O

O O
"
B R' + R OH R" + R'OH
R O R O

O O
R' + R"NH2 R" '
+ R OH
C R N
R O
H

O O
D R' + R"SH
R S
R" '
+ R OH
R O
H
Lipaze- serin hidrolaze
O O
ester hydrolysis (A), O O
R' + R' +
alcoholysis (B), E R O "R OH "R O R OH
aminolysis (C),
thiolysis (D), O O O O
acidolysis (E), R' + R"' R' + R"'
F R O "R O "R O R O
interesterification (F)
 Situsul activ al chimotripsinei

His 57
H2C -serine residue (Ser105) is activated by a
Asp 102 O H N N Ser 195 hydrogen bond to histidine (His224). The result
CH2 C
O
H O CH2
of this charge relay eliberation is potentate by
Ser nucleophilicity which is very important for
catlytic activation.

His 57
Asp 102 O H2C
Ser 195
CH2 C
O H N N H O CH2

Reeaua de eliberare de sarcin a chimotripsinei

 His
H2C

Cataliza chimotripsinei i a altor Asp O H N N Ser

serinproteinaze
CH2 C H O CH2
O
O
are loc n cinci etape: R1
CH
O
C
CH
CH

- n chimotripsin, substratul este R2

meninut ntr-un
anumit loc, cu ajutorul interaciunilor ce H2C
His

se Asp O N N H
formeaz ntre substrat i enzim; Ser
CH2 C O CH2
O H

O
-Hidroxilul nucleofilic al serinei Sr 195 R1
CH
O
C
CH
CH

atac C carbonilic din legtura peptidic, R2

R1 CH OH
formnd un H2C
His

intermediar tetraedric; Asp O H N N Ser

CH2 C O CH2
H
O
- n cea de-a treia etap, are loc o O
H O

cataliz general C
CH
CH

R2
acid, legtura peptidic fiind rupt,
gruparea fugacee prelund un proton de His

la ionul imidazol al restului de histidin, H2C

His 57; Asp O N N H Ser

CH2 C O CH2
O H

-Poriunea acil din legtura peptidic O

C
O

CH
CH

rmne legat de enzim ca intermediar H2C

His
H R2

acil-enzim; Asp H N N
O Ser
CH2 C H O CH2
O
-n urmtoarea etap, apa hidrolizeaz O
H O C CH
intermediarul acil-enzim, genernd un CH
R2
alt intermediar tetraedric;
Practical applications of lipases in the resolution of racemates
O
R O Lipase R R
+ toluene + +
OH O O OH

O
(RS)-alcohol isopropenyl acetate (R)-ester (S)-alcohol

HO OH

O
OH OH OH
ee>99% ee>99% ee>99% ee>99% ee>94%
E= 93 E= 287 E>300 E=4 E= 26
OH
OH OH OH
H3CO

ee>99% ee>99% ee>99% ee>99%

E= 141 E= 78 E= 16 E= 23
OH OH F F
F

HO OH
ee>99% ee>99% ee>99% ee>99%
E= 9 E= 10 E= 42 E= 29

Lipase-catalyzed transesterification of secondary alcohols using isopropenyl acetate

as acyl donor in toluene

Ghanem, A., Aboul-Enein H. Y., Tetrahedron: Asymmetry, 2004, 15, 33313351;

 Selected examples of the kinetic resolution of secondary alcohols

OH

OH OH OH O
OH
R O N3 O S O
Ar Ar R1 R O
O O
ee= 98.6% ee= 62-96% ee= 85%

O ee>98% ee>99%
O OH
N
N O R O
OH HO OH
O R
Lipase Lipase
PSL
ee>99% vinyl acetate vinyl acetate
vinyl acetate
ee= 32-86% ee= 60-92%
ee: n.d
OR O
RO OR OR
OR
O
O O O
R= H R= H R= H R= H
ee: 64-93% ee: 25-94% ee: 28-99% R= H
ee: 35-85% ee: 24-71%
R=Ac R=Ac R=Ac R=Ac
ee> 96% ee= 45-99% ee= 3->99% R=Ac
ee= 73-93% ee= 15-83%

Paizs, C.; Tosa, M.; Bodai, V.; Szakacs, G.; Kmecz, I.; Simandi, B.; Majdik, C.; Novak, L.; Irimie, F.D.; Poppe, L. Tetrahedron: Asymmetry 2003, 14, 19431949; Pc
K. B.; Loupy, A.; Plenkiewicz, J.; Blanco, L. Tetrahedron: Asymmetry 2000, 11, 27192732; Wielechowska, M.; Plenkiewicz, J. Tetrahedron: Asymmetry 2003, 14
Acherar, S.; Audran, G.; Vanthuyne, N.; Monti, H. Tetrahedron: Asymmetry 2003, 14, 24132418; Nascimento, M. G.; Zanotto, S. P.; Melegari, S. P.; Fernandes,
Tetrahedron: Asymmetry, 2003, 14, 31113115; Vorlova, S.; Bornscheuer, U. T.; Gatfield, I.; Hilmer, J.-M.; Bertram, H.-J.; Schmid, R. D. Adv. Synth. Catal. 2002
Buchalska, E.; Plenkiewicz, J., J. Mol.Catal. B: Enzym. 2001, 11, 255263; Joly, S.; Nair, S. M. J. Mol. Catal. B: Enzym. 2003, 22, 151160; Baumann, M.; Hauer
Bornscheuer, U. T. Tetrahedron: Asymmetry 2000, 11, 47814790; Bierstedt, A.; Stoelting, J.; Froehlich, R.; Metz, P. Tetahedron: Asymmetry 2001, 12, 33993
 The lipase-catalyzed enantioselective acylation of a chiral amine .

Gerard Gil, J. Org. Chem., 2007, 72 (18), pp 69186923

 Lipase-catalyzed enantioselective acetylation of bicyclic 1 -heteroaryl primary amines

Kinetic resolution of azido acetate 123

Ashraf Ghanem, Chirality, 17, 115 (2005);
 APPLICATION OF LIPASES IN INDUSTRY

Some of the biocatalytic steps using lipase developed at BASF: Lipase-catalyzed kinetic
resolution of (a) phenyl ethanol 18 using succinic anhydride, (b) secondary amine 131 using ethyl
methoxyacetate as acyl donor;

Enantioselective hydrolysis of racemic acetamide (R,S)-133 developed at Bayer

Ashraf Ghanem, Chirality, 17, 115 (2005);

 Enantioselective hydrolysis of racemic glycidylbutyrate (135) developed at Andeno-DSM

Lipase-catalyzed hydrolysis of the terminal vinyl ester in taxol 2V-vinyladipate 137

Ashraf Ghanem, Chirality,17, 115 (2005);

 Acylase I
O O O
R R R
OH acylase OH OH
+
HN pH 7.0-7.3 NH2 HN
20-37oC
O O
R = H3CSC2H4, HSCH2,H3CSCH2, H3CS(O)2C2H4, CH3, C2H5, (H3C)2CH, C3H7, C4H9

Zn2+ Zn2+

Amino O O
acylase I C CH2 R1 COO C CH2 R1 COO
R N R N
O
H 1 H H O
H
H O C Glu
O
H
O
C Glu
O 2

Zn2+ Zn2+

O O
C 3
C CH2 R1 COO
R O + H CH2 R1 COO
N R N
O
H H
H

B H B

Mechanism of action of a metalloprotease

 Lipase catalyzed kinetic resolution of the
racemic 1-heteroaryl derivatives

Enzymatic reactions
O

OH lipase OH O
+
R vinyl acetate R R

5a-d (S)-5a-d (R)-6a-d

lipase
EtOH

N OH
R R
S S S O
a b c d (R)-5a-d
Chemical synthesis
Cl O
O
I. II. III.
S S S S S
4b 4c O
O O O

IV. V. VI. VII.

O OH O
OH O O O
O O
1 2 3 4d

I.BuLi, -78 C, 1 h, 2) DMF, -10 C, 2 h; ; II.CH2O/HCl, 60oC;III. urotropine/CH3Cl, toC

IV.ClCH2COOEt, K2CO3 / Acetone; V. K2CO3 / H2O; VI. Ac2O / AcO-Na+;
VII. SeO2 / Dioxane, reflux;

Synthesis of heteroaryl adehydes

N OH O OH
I. N II.

S S R R
4b-d 5b-d
5a
O
OH III. O
R R
5a-d 6a-d

N
R
S S S O
a b c d

I. 1) BuLi, -78 C, 1 h, 2) CH3CHO, -10 C; II. CH3MgI/ether;

III. acetic anhydride, Et3N, DMAP, CH2Cl

Synthesis of racemic heteroaryl ethanols and corresponding acetates

Substrate rac-2a
rac-2a rac-2b
rac-2b rac-2c
rac-2c rac-2d
rac-2d

Time Time Time Time

( ( ( (
Lipase ee/s % ee/p % c% ee/s % ee/p % c% ee/s % ee/p % c% ee/s % ee/p % c%
h h h h
) ) ) )

AK 6 >99.5 >99.5 50.0 24 >99.5 >99.5 50.0 16 >99.5 >99.5 50 24 >99.5 >99.5 50.0

Novozyme 435 72 >99.5 >99.5 50.0 24 98.0 97.5 50.2 16 >99.5 97.7 50.6 24 96.6 96.3 50.0

AY 19 84.3 77.6 52.0 46 60.4 89.5 40.3 40 21.8 82.0 21 46 32.6 82.8 28.2

CrL 6 86.8 16.5 84.0 24 33.8 >99.5 25.3 40 7.2 >99.5 6.7 24 12.3 72.7 14.5

TL-IM 26 65.3 11.1 85.4 46 25.2 90.3 21.9 40 18.3 >99.5 15.5 46 10.7 >99.5 9.7

CCL 5 89.9 53.0 62.9 46 55.0 89.0 38.2 48 66.3 99.5 40 46 35.7 1.7 95

PPL 26 85.7 25.7 76.9 46 11.0 >99.5 9.9 48 6.7 97.9 6.4 46 6.1 >99.5 5.8

PS 24 >99.5 93.2 51.8 46 94 85.8 52.3 48 3 10.3 22.9 46 26.9 >99.5 21.2

Mucor
javanic
us 44 >99.5 2.7 97.4 24 7.6 >99.5 7.0 24 39.8 95.5 29.4 24 0.95 >99.5 0.9

Rhiz arrheus
inverse
selectivi
ty 44 50.0 13.6 78.6 120 9.4 6.7 58.55 40 6.8 66.6 9.2 120 2.3 93.3 2.5

F inverse
selectivi
ty 26 89.3 45.9 66.0 24 32.0 92.7 25.7 21 >99.5 >99.5 50.0 24 14 22.7 38.3

3b 38 >99.5 >99.5 50.0 15 >99.5 >99.5 50.0 21 42.6 >99.5 29.9 22 56.7 >99.5 36.2

9 38 >99.5 >99.5 50.0 15 84.4 >99.5 45.8 21 30.5 >99.5 23.4 22 13.6 >99.5 11.9

23 38 32.3 >99.5 24.4 15 56.6 >99.5 36.1 21 13.9 >99.5 12.2 22 11.1 >99.5 10.0

97 38 58.3 >99.5 36.8 15 63.3 >99.5 38.8 21 10.4 >99.5 9.4 22 6.7 95.7 6.5

101 38 29.1 >99.5 22.5 15 41.7 >99.5 29.4 21 20.9 >99.5 17.3 22 0.6 >99.5 0.5

117 38 48.4 85.2 36.2 15 80.6 >99.5 44.6 21 1.8 >99.5 1.8 22 2.8 >99.5 2.7

158 inverse
selectivi
ty 38 19.8 98.6 16.8 15 21 2.5 >99.5 2.4 22 15.8 60.4 20.7

171 inverse
selectivi
ty 38 20.5 76.5 21.1 15 83.9 35.0 70.6 16 >99.5 >99.5 50 22 70.9 63.2 52.9
Preparative scale kinetic resolution of the racemic heteroaryletahanols
catalyzed by Novozym 435

ee Yield Yield ee
E C
(%) (%) (%) (%)
N OH N OAc

S 97.2 44 >200 50.4 45 98.0 S

OH OAc

S 99.1 45 200 49.8 46 99.3 S

AcO
HO

96.5 46 >200 42.2 43 97.8

S
S
HO AcO

97.8 43 >100 48.1 45 96.7

O
O
Lipase-catalyzed alcoholysis of the racemic acetates
O O

O O OH
lipase
+ Ar
Ar R-OH Ar
rac (S) (R)

R=Me,Et,Pr,Bu

Ethanolysis with Novozym 435

ee (S)-acetate ee (R)-ethanols
Heterocycle c [%] E
[%] [%]
N

S
51.6 99.3 97.5 >100

S 49.7 94.6 97.5 >100

S 22.1 21.8 99.3 >100

O 18.1 21.8 95.6 55

 Rezolutia cinetica dinamica(DKR)

Rezolutia cinetica dinamica este in principiu un proces de rezolutie cinetica in care

enantiomerul netransformat este racemizat in situ.
- Cu aceasta tehnica se poate ajunge la o converisie de 100%

RS + SS RP + SS

racemization

R COOH R COOH

acilaza R COOH
HN R' HN R' +
NH2
+
O H /t O
racemizare D L
 DKR of -aminonitriles to form chiral -amino acids.

K. Yasukawa, R. Hasemi and Y. Asano, Adv. Synth. Catal., 2011, 353, 2328

DKR of aromatic alcohols using different acyl donors.

G. Xu, Y. Chen. J. Wu, Y. Cheng and L. Yang, Tetrahedron: Asymmetry, 2011, 22, 1373
DKR with racemization catalyst I and CALB (Novozyme 435)
Dynamic kinetic resolution of oxazolone
derivatives

O
O

R
R OR2
O rapid
R2-OH N O
N
H
R1
R1

rapid racemizare

O
O

R
R lent OR2
O

R2-OH N O
N H
R1
R1
 Ph H O Ph H O Ph H O

N O N O N O O O
COOBu
Ph Ph Ph A. niger lipase lipase(Mucor miehei)
+
PPL
pKa 8.9/9.5
N O n-BuOH
N O H N Ph
Ph
Ph O
Ph Ph O
O
Ph N COOH L-t-Leu
Ph N COOH H
H
ee= 99% ee= 99%
in situ racemization
Turner N.J., Winterman J.R., McCague R., Parrat J.S., Taylor S.J.C.,
Gu R.L., Lee I.S., Sih C.J., Tetrahedron Lett., 1992, 33, 1953 Tetrahedron Lett.,1995, 36(7), 1113-115;
X
Ph
HN Ph
N NuOH
X O
Enzyme R Nu
R
O O

R X Enzyme NuOH Solvent Yield(%) ee(%)

Ph-CH2- O Novozyme MeOH MeOH 81 95(S)

Ph-CH2- O Novozyme EtOH MeOH 82 97(S)

Ph-CH2- O Novozyme MeOH CH3CN 88 98(S)

i-Bu O Novozyme MeOH CH3CN 96 97(S)

i-Bu O Lipozyme n-BuOH Toluene 67 99.5(S)

Ph O Lipase P-30 MeOH MTBE 46 75(S)

Ph-CH2- O Lipase P-30 MeOH MTBE 80 78(S)

Ph-CH2- O Liapse PL H2 O Buffer pH 7.6 >95 >99(S)

Ph-CH2- O Lipase AP H2 O Buffer pH 7.6 >95 >99(R)

CH3SCH2CH2 S Chymotrypsin H2 O Buffer pH 6.8 97 94

n-Bu S Prozyme 6 H2 O Buffer pH 6.8 86 99(S)

Brown S.A., Parker M.C., Turner N.J., Tetrahedron Asymmetry, 2000, 11, 1687-1690
 Chemoenzymatic preparation of enantiopure
L-benzofuranyl- and L- benzo[b]thiophenyl alani

R R1 OH COOR1
O R
N fast
NHCOCH3

racemic fast racemization

DKR mixture
O
R R1 OH COOR1
R
O
N slow NHCOCH3

O
COOH DCC COOR
R R ROH
CH2Cl2 O R
NHCOCH3 N lipase
solvent NHCOCH3

100% theoretic yield

S S O O
 Experimental part
hemoenzymatic preparation of enantiopure L-benzofuranyl- and L- benzo[b]thiophenyl alanines

O COOH COOH COOH

R AcylaseI R R
R O + b
NHCOCH3 pH 7-8 NH2 NHCOCH3
N
9a-d L-12a-d D-9a-d
D-11a-d O S
a b
O
I. COOPr II. COOH
R O R R
a N NHCOCH3 NHCOCH3
O S
c d 11a-d L-10a-d (ee 81-87%) L-9a-d (ee 81-87%)

c III.
O
COOAlk COOH
R O
Lipase
R I. Propanol, Novozyme 435 / Dioxane; R
N II. Na2CO3, H2O, reflux;
Alcohol NHCOCH3 NH2
III. Acylase I, pH 7-8.
L-11a-d L-10a-d
L-12a-d (ee>99%)

Enantioselective synthesis of L -heteroaryl alanines and their derivatives

a. DKR of the lipase mediated alcoholiysis of oxazolones (rac)-11a-d;
b. enantiomer selective hydrolysis of (rac)- 9a-d;
c. chemoenzymatic synthesis of enantiopure L-12a-d
Enzymatic synthesis
O
Lipase COOAlk
R O R
N Alcohol NHCOCH3
L-10a-d
rac-11a-d

O S
a b

O S
c d

-Several enzymes were tested. All lipases, excepting Lipase F, showed the same enantiopreference in their alc
although the behaviour of lipases greatly differed. Lipases AY, PS, CcL and CrL were catalytically inactive, lipa
(e.e. 13-17%), lipase F (e.e. 5- 7%) and Lipozyme TL IM (e.e. 7-11 %) gave moderate selectivity and reactivity
whereas Novozyme 435 showed acceptable properties.

Entry Type of the e.e. for L-2-amino-3-(heteroaryl)propanoic esters

ester (%)a, b
L-10a L-10b L-10c L-10d

1 Methyl ester 3 3 3 3
2 Ethyl ester 67 65 65 73
3 Propyl ester 71 71 69 79
4 Buthyl ester 67 69 67 75

a in neat alcohols

b total conversion after 6 hours

 O
COOPr
R O
Novozym 435 R
N 2 eq. PrOH, solvent NHCOCH3
L-10a-d
rac-11a-d

O S
a b

O S
c d

Entry Solvent E.e. for L-2-amino-3-(heteroaryl)propanoic acid propyl

esters (%)
L-11a L-11b L-11c L-11d

1 Dioxan 75 79 83 86
2 Dichloromethane 39 37 42 54
3 Toluene 69 59 69 79
4 Acetonitrile 61 55 73 83
5 Tetrahydrofuran 71 47 81 83
6 Diethylether 67 71 59 73

Enantiomeric excess for DKR of 4-((heteroaryl)methyl)-2-methyloxazol-5(4H)-ones 11a-d in organic solv

 ELUTION DIAGRAM:
for the enzymatic DKR
O
O

S
O O
O O

NHCOCH3 NHCOCH3

ELUTION DIAGRAM:
S S

of the mixture of the racemic of starting material

and racemic product for the lipase catalysed DKR
after2h
O O
O O
O O
N NHCOCH3
NHCOCH3

S S
S

O O
O O
O O
N NHCOCH3
NHCOCH3

S S
S

after24h

O
O O
O
NHCOCH3
NHCOCH3
S
S

after48h
O
O

S
 COOH COOH COOH
R Acylase I R R
+
NHCOCH3 pH 7-8 NH2 NHCOCH3
9a-d L-12a-d D-9a-d

O S
a b

O S
c d

Entry Substrate c (%) ee% L-12 ee% D-9

1 9a 50.0 >99.5 >99.5

2 9b 50.0 >99.5 >99.5

3 9c 50.0 >99.5 >99.5

4 9d 50.0 >99.5 >99.5

Enantiomeric excess for enantiomer selective hydrolysis of (rac)- 9a-d catalyzed by Acylase I
Chemoenzymatic synthesis of enantiopure L-12a-d
O
COOH I. II. COOPr III. COOH
R R O R R
NHCOCH3 N NHCOCH3 NHCOCH3
rac-9a-d rac-11a-d L-10a-d (ee 81-87%) L-9a-d (ee 81-87%)

IV.

COOH
O S R
a b NH2
L-12a-d (ee>99%)
I. DCC/CH2Cl2, 0oC; II. Propanol, Novozyme 435 / Dioxane;
O S III. Na2CO3, H2O, reflux; IV. Acylase I, pH 7-8.
c d

Substrate Product a Yield []D20 (10 mgmL-1)

(%)b

-14.5, CH3COOH
(rac)-9a L-12a 76
(-14.5, CH3COOH, 20C)

-23.8, CH3COOH
(rac)-9b L-12b 79
(-23.8, CH3COOH, 20C)

(rac)-9c L-12c 83 -8.0, CH3COOH

-26.0, CH3COOH
(rac)-9d L-12d 85
(-9.17, 0.1 N HCl, 20C)
a e.e. 99% for all the compounds
b yields are given for the isolated compounds
 COOC3H7 Na CO H O COOH AcylaseI COOH
R 2 3, 2 R
reflux R
NHCOCH3 NHCOCH3 pH 7-8
NH2
L L L
ee =75-86% ee >99%
Elution diagram of the isolated enantiopure amino acid
after Acylase I mediated hydrolysis

Yield (%)
79
S

85
S

83
O

O 76

ee 99 %