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An enzyme-catalyzed synthesis of natural α-tocopherol
095%4166/93w&Ml0 PergalnollprrrsLtd
Tandtubt:Asyw~ Vol. 4, No. 9. pp. 1961-1964.1993 %%tdinGRatBlitain
Eisaku Mizuguchi, Masumi Taktmoto, and Kszuo Achiwa* annsceutieal seifnces,udversjtyof schoOlOfPh
z5hhudw
52-I Y&I, ShiEuoiu 422, Japan
(Received in Japan 16 Juno 1993)
The a-tocopherol Mamin E) is known as a potent and safe, lipid-soluble antioxidant. Recently, tocopherol and super oxide dismtase@OD) and other antioxidants are attracting attention as scavengers of super oxide seri&.
As tocoptil
is being used for various purpares, in the future, demand will more increase. Presently,
the majopity of tocopherol is being used as a mixture of its eight isomw, which weie mpofted to show different biological activities. Tocopherof with the S configuration at the 2-position of its chroman ring is weak in its biological activities2. Rreviousiy, d&ocopheroljWS,4’R,8’R)-I] has been resolved into d- and I-tocopherol [@I - and W-11 by Karrer” and Robeson4et al . using 3-bromocamphor-&sulfonate or piperazine, reqectively. However, their optical resolution is not efficient and of less utiiity val&. After investigation, we found an effective synthesis of natural a-tocopherol using the enzyme-catalyzed kinetic resolution and the conversion of the unnatural enantiomer to a-tocopherol by its racemization and repeated enzyme-catalyzed hydrolysis.
First, enzymatic hydrolysis of dl-tooopherol
acetate and benzoate were examined, but these reactions were
unsuccessful. Next, we chose to& acetate[W)-71 as less sterically hindered substrate. Its ester site and stexeogenic carbon atom of the chromsn 2-position were more able to interact with enzymes. The O-7 was synthesized as shown in Scheme 1. Table 1 shows the results of its enzyme-catalyzed hydrolysis. The hydrolysis of @S&7 with lipaseAY6 gave (R)-77 in extremely high optical yield f>We), which was converted to natural a-toqhm1 IGR)-118 WHKBUC r-am&&n by dial acco&ng to fijima’s method? It is an interest@ result that t&S)=7is hydrolyzed with high enantioaelectivity by lipaseAY, in spite of the reaction site being fairly remote from the steorogenic castxm atom of its chroman ring. To out knowledge, no e.nzyme-cata@zedenantioselective hydrolysis of this tyqe of substrate haa baeo rqxmedt”. l%l
1962
E. kflZUCiUCH1
Scheme 1
OH
al.
Enzymatic Synthesis of d-a-Tccopherol a
2
et
-
75%
R
w 3
b 94%
O
4
I
OH 5
(RS)-6
Enzyme
(RS)-7
\
*
H*O/lPE
Reagents; a KMn04/acetone 6 vinyl bromide+B&i/Et20 c BFrEtaOQO dAcCI, Et$VTHF e CH20,HsB04,HzlPd-CI(CHsO)~B f AcCl,Et3NTTHF,Na2Cr2Cr/AcDH g NaOWEtOH h NaBH4, then Ha/Pd-CIMeOH
Table 1 Lipase-cafalyzed Kinefic Resolution of To&acetate
RS-7 a
W6
w7
mY-7
Enzyme”
1
lipase AY
1
32
>99
60
36
2
CHE
46
42
76
50
55
3
liise AH
48
78
2.5
16
12
4
lipase PS
48
100
0
0
0
5
lipase M
46
100
0
0
0
a All reactions
Time(h) C.Y. (%I b
O.Y. (%ee)’
C.Y. (%I b
0-Y. (%ee) ’
Entry
were carried out by stlmng a mixture 01 substrate(100n-g )lipase(lOOmg~and IPE saturated with waler at 25%. b Isolated yield c Opbcal yields 017 were determIned by HPLC analyses using a column packed with Ch~ralcel OD-i-l (2-propanol/hexane),and optical yields of 6 were detefmvNd after conversK)n to 7.
Synthesis of natural a-tocopherol
1963
Scheme 2
H20/lPE
w-10 kc-tocopherol
W-11
Reagents, a pyrrolidine/toluene b AcCI,Et$WHF c NaB&.thenHzIPd-CIMeOH d CH20, H3B0,,H~Pd-C/(CH30)3B e EtONa/EtOH
Table 2 Lipase-catalyzed Kinetic Resolution of RS- 10 a
w-8
(R)_l 0
ms~-lo Entry
Enzyme’
Time(h)
1
IipaseAH
2
13
42
87
31
2
IipaseAK
8
36
18
64
39
3
IipaseM
24
40
74
55
40
4
IipaseAY
0.7
18
9.7
73
22
5
F-AP15
72
53
13
40
41
C.Y.(%OJ.Y.(%eeIc
C.Y.(%)QD.Y.(%ee)c
a All reactK)ns were carried out by siimng a mixture of substrate(l00mg)llpase(lOOmg)afld IPE saturated wlh water al 25%. b lsohted yield. c Optical yields of 10 were determtned by HPLC analyses using a column packed wrfh Chlralcel OD-H (2-propanol/hexane).and optical yields 8 were determlned after converslon to 10.
E. IvItzuouc~~et al.
1964
Acetylation of the other enantiomer [O-6] and subsequent oxidation with Na2Cr2@ gave 4-oxotccol acetate [(5’)-lo], which was easily racemized with NaOEt. Reduction of the @5)-)-sthus obtained with NaBH4 and subsequent hydrogenation with W-C gave (KY)-6. This (RT)-6 was then employed as a substrate of enzyme-catalyzed hydrolysis. Alternatively, t&!&l0 was synthesized directly from 2,Sdihydroxyacetophenone (9) as shown in Scheme 2 and similarly submitted to enzyme-catalyzed hydrolysis. Table 2 shows the results. The (R)-10” obtained by enzyme-catalyzed kinetic resolution was reduced to f.@-tocol by NaBHqreduction and W-C catalyzed hydrogenation, which was then methylated to give natural a-tocopherol [@)-I]*. The unnatural enantiomer [Gs)-81 was converted to the substrate [O-101
for the enzyme-catalyzed hydrolysis by racemization and
subsequent acetylation.
REFERENCES
AND NOTES
1.
Niki,B.;KonnuoE.; “OxygenRadicaLsin Biology and hfedicine”,(Simic.M.G.;Taylor.K.A.;WardJ.F.;
2.
(a)Ames.S.R.;Ludwig.M.I.;Nelm~D.R.;R0beson.C.D.;Biochemistry,l9632,188.
c.voll Somltageds.)
(b)MacMin.LJ.;Gsbriebriel.E.; B&M.;
J.Nutr. 1982.112.1437.
Karrer.P.;Koenig.H.;Ringier.B.H.;Sslomon.H.;Helv.Chem.Acta,1939.22,1139. Robeson.C.D.;Nelan.D.R.: J.Am.Chem.Soc.1962,84.3196.USP.3.153.04&1964. Schudel.P.;Mayer.H.;Matzged.;Ruegg.R.;Isler.O.;Helv.Chem.Acta.l%3.46,333, Enymes were kindly supplied by Amen0 Flumxeutical CHE(Cholesed
Co.,Ltd. @sseAY(Candida fugosa)
es&rase).lipaseAH(sp.)~ipssePS(Pse~o~~sp.),tipaseM(Mucorjavanicus)
lipaseAK(?%e&monas sp.),F-APlWWzopus javankud. 7.
(R)-?: [a]D2’ +4.1 (c 0.6 EtGH). lH-NMR(CDC13) 6: 0.80-0.88(12H.m).1.05-1.65 (24H.m). 1.651.91(2H,m), 2.25(3H.s),2.70(2H,tJ=4.4Hz), 6.53-7.67(3H.m).
8.
identificationwss csrriedout by comparisonof HPLC analysisusinga cohnmtpsckedwitb Daicel Chimkel OD-H(bex&-pq.xmol=2~0/1)
with an authentic d-a-tocopbeml.
(R)-l:[a]Dm +0.68 (c 0.8 EtOH). [lit.5 +0.75(EtOH), authenticd-a-tocopheml+O.70(c 1.0 EtOH)]. lo-NMR (CDC13) 8: 0.82-0.88(12H,m),1.07-1.60(24H,m),1.72-1.85(2H,m),2.10-2.18(9H,m), 2.60(2H.t.l=2Hz). 4.18(1H.s).MASSzn/z(M+)430. 9.
Kijima.S.:Nonska.A.; J’P.60-4183(1985).
10.
Pmvioosly,wehave reportedsnothertype of substrate. of which msctionsite a little remotefnnn the stereogeniccarbonatom.(a)Mumts.M.;AcbiwaK.; Tetmhedfvn Left, 1991.32.6763. (b)Ebike.H.;Temo.Y.;A&iwa.K.; TetrahedronLett,1991.32.5805.
11.
(R)-l(:[a]#’
+9.02 (c 0.6 EtOH) ‘H-NMR (CDC13) 6: 0.83-0.90(12H.m),LOO-l.80
(24H_m),2.25(3H.s),2.70(2H.ddJ=17Hz).6.91-7.54(3Hm).
Tandtubt:Asyw~ Vol. 4, No. 9. pp. 1961-1964.1993 %%tdinGRatBlitain
Eisaku Mizuguchi, Masumi Taktmoto, and Kszuo Achiwa* annsceutieal seifnces,udversjtyof schoOlOfPh
z5hhudw
52-I Y&I, ShiEuoiu 422, Japan
(Received in Japan 16 Juno 1993)
The a-tocopherol Mamin E) is known as a potent and safe, lipid-soluble antioxidant. Recently, tocopherol and super oxide dismtase@OD) and other antioxidants are attracting attention as scavengers of super oxide seri&.
As tocoptil
is being used for various purpares, in the future, demand will more increase. Presently,
the majopity of tocopherol is being used as a mixture of its eight isomw, which weie mpofted to show different biological activities. Tocopherof with the S configuration at the 2-position of its chroman ring is weak in its biological activities2. Rreviousiy, d&ocopheroljWS,4’R,8’R)-I] has been resolved into d- and I-tocopherol [@I - and W-11 by Karrer” and Robeson4et al . using 3-bromocamphor-&sulfonate or piperazine, reqectively. However, their optical resolution is not efficient and of less utiiity val&. After investigation, we found an effective synthesis of natural a-tocopherol using the enzyme-catalyzed kinetic resolution and the conversion of the unnatural enantiomer to a-tocopherol by its racemization and repeated enzyme-catalyzed hydrolysis.
First, enzymatic hydrolysis of dl-tooopherol
acetate and benzoate were examined, but these reactions were
unsuccessful. Next, we chose to& acetate[W)-71 as less sterically hindered substrate. Its ester site and stexeogenic carbon atom of the chromsn 2-position were more able to interact with enzymes. The O-7 was synthesized as shown in Scheme 1. Table 1 shows the results of its enzyme-catalyzed hydrolysis. The hydrolysis of @S&7 with lipaseAY6 gave (R)-77 in extremely high optical yield f>We), which was converted to natural a-toqhm1 IGR)-118 WHKBUC r-am&&n by dial acco&ng to fijima’s method? It is an interest@ result that t&S)=7is hydrolyzed with high enantioaelectivity by lipaseAY, in spite of the reaction site being fairly remote from the steorogenic castxm atom of its chroman ring. To out knowledge, no e.nzyme-cata@zedenantioselective hydrolysis of this tyqe of substrate haa baeo rqxmedt”. l%l
1962
E. kflZUCiUCH1
Scheme 1
OH
al.
Enzymatic Synthesis of d-a-Tccopherol a
2
et
-
75%
R
w 3
b 94%
O
4
I
OH 5
(RS)-6
Enzyme
(RS)-7
\
*
H*O/lPE
Reagents; a KMn04/acetone 6 vinyl bromide+B&i/Et20 c BFrEtaOQO dAcCI, Et$VTHF e CH20,HsB04,HzlPd-CI(CHsO)~B f AcCl,Et3NTTHF,Na2Cr2Cr/AcDH g NaOWEtOH h NaBH4, then Ha/Pd-CIMeOH
Table 1 Lipase-cafalyzed Kinefic Resolution of To&acetate
RS-7 a
W6
w7
mY-7
Enzyme”
1
lipase AY
1
32
>99
60
36
2
CHE
46
42
76
50
55
3
liise AH
48
78
2.5
16
12
4
lipase PS
48
100
0
0
0
5
lipase M
46
100
0
0
0
a All reactions
Time(h) C.Y. (%I b
O.Y. (%ee)’
C.Y. (%I b
0-Y. (%ee) ’
Entry
were carried out by stlmng a mixture 01 substrate(100n-g )lipase(lOOmg~and IPE saturated with waler at 25%. b Isolated yield c Opbcal yields 017 were determIned by HPLC analyses using a column packed with Ch~ralcel OD-i-l (2-propanol/hexane),and optical yields of 6 were detefmvNd after conversK)n to 7.
Synthesis of natural a-tocopherol
1963
Scheme 2
H20/lPE
w-10 kc-tocopherol
W-11
Reagents, a pyrrolidine/toluene b AcCI,Et$WHF c NaB&.thenHzIPd-CIMeOH d CH20, H3B0,,H~Pd-C/(CH30)3B e EtONa/EtOH
Table 2 Lipase-catalyzed Kinetic Resolution of RS- 10 a
w-8
(R)_l 0
ms~-lo Entry
Enzyme’
Time(h)
1
IipaseAH
2
13
42
87
31
2
IipaseAK
8
36
18
64
39
3
IipaseM
24
40
74
55
40
4
IipaseAY
0.7
18
9.7
73
22
5
F-AP15
72
53
13
40
41
C.Y.(%OJ.Y.(%eeIc
C.Y.(%)QD.Y.(%ee)c
a All reactK)ns were carried out by siimng a mixture of substrate(l00mg)llpase(lOOmg)afld IPE saturated wlh water al 25%. b lsohted yield. c Optical yields of 10 were determtned by HPLC analyses using a column packed wrfh Chlralcel OD-H (2-propanol/hexane).and optical yields 8 were determlned after converslon to 10.
E. IvItzuouc~~et al.
1964
Acetylation of the other enantiomer [O-6] and subsequent oxidation with Na2Cr2@ gave 4-oxotccol acetate [(5’)-lo], which was easily racemized with NaOEt. Reduction of the @5)-)-sthus obtained with NaBH4 and subsequent hydrogenation with W-C gave (KY)-6. This (RT)-6 was then employed as a substrate of enzyme-catalyzed hydrolysis. Alternatively, t&!&l0 was synthesized directly from 2,Sdihydroxyacetophenone (9) as shown in Scheme 2 and similarly submitted to enzyme-catalyzed hydrolysis. Table 2 shows the results. The (R)-10” obtained by enzyme-catalyzed kinetic resolution was reduced to f.@-tocol by NaBHqreduction and W-C catalyzed hydrogenation, which was then methylated to give natural a-tocopherol [@)-I]*. The unnatural enantiomer [Gs)-81 was converted to the substrate [O-101
for the enzyme-catalyzed hydrolysis by racemization and
subsequent acetylation.
REFERENCES
AND NOTES
1.
Niki,B.;KonnuoE.; “OxygenRadicaLsin Biology and hfedicine”,(Simic.M.G.;Taylor.K.A.;WardJ.F.;
2.
(a)Ames.S.R.;Ludwig.M.I.;Nelm~D.R.;R0beson.C.D.;Biochemistry,l9632,188.
c.voll Somltageds.)
(b)MacMin.LJ.;Gsbriebriel.E.; B&M.;
J.Nutr. 1982.112.1437.
Karrer.P.;Koenig.H.;Ringier.B.H.;Sslomon.H.;Helv.Chem.Acta,1939.22,1139. Robeson.C.D.;Nelan.D.R.: J.Am.Chem.Soc.1962,84.3196.USP.3.153.04&1964. Schudel.P.;Mayer.H.;Matzged.;Ruegg.R.;Isler.O.;Helv.Chem.Acta.l%3.46,333, Enymes were kindly supplied by Amen0 Flumxeutical CHE(Cholesed
Co.,Ltd. @sseAY(Candida fugosa)
es&rase).lipaseAH(sp.)~ipssePS(Pse~o~~sp.),tipaseM(Mucorjavanicus)
lipaseAK(?%e&monas sp.),F-APlWWzopus javankud. 7.
(R)-?: [a]D2’ +4.1 (c 0.6 EtGH). lH-NMR(CDC13) 6: 0.80-0.88(12H.m).1.05-1.65 (24H.m). 1.651.91(2H,m), 2.25(3H.s),2.70(2H,tJ=4.4Hz), 6.53-7.67(3H.m).
8.
identificationwss csrriedout by comparisonof HPLC analysisusinga cohnmtpsckedwitb Daicel Chimkel OD-H(bex&-pq.xmol=2~0/1)
with an authentic d-a-tocopbeml.
(R)-l:[a]Dm +0.68 (c 0.8 EtOH). [lit.5 +0.75(EtOH), authenticd-a-tocopheml+O.70(c 1.0 EtOH)]. lo-NMR (CDC13) 8: 0.82-0.88(12H,m),1.07-1.60(24H,m),1.72-1.85(2H,m),2.10-2.18(9H,m), 2.60(2H.t.l=2Hz). 4.18(1H.s).MASSzn/z(M+)430. 9.
Kijima.S.:Nonska.A.; J’P.60-4183(1985).
10.
Pmvioosly,wehave reportedsnothertype of substrate. of which msctionsite a little remotefnnn the stereogeniccarbonatom.(a)Mumts.M.;AcbiwaK.; Tetmhedfvn Left, 1991.32.6763. (b)Ebike.H.;Temo.Y.;A&iwa.K.; TetrahedronLett,1991.32.5805.
11.
(R)-l(:[a]#’
+9.02 (c 0.6 EtOH) ‘H-NMR (CDC13) 6: 0.83-0.90(12H.m),LOO-l.80
(24H_m),2.25(3H.s),2.70(2H.ddJ=17Hz).6.91-7.54(3Hm).