I HC QUC GIA H CH MINH

TRNG I HC BCH KHOA

KHOA K THUT HA HC
B MN CNG NGH SINH HC

n chuyn ngnh

CY THNG
V HP CHT TAXOL

CBHD: TS. L TH THY TIN

SVTH: NGUYN H HONG ANH
MSSV: 60700056
B MN: CNG NGH SINH HC

Tp. HCM, thng 06 nm 2011

LI CM N
Trong qu trnh thc hin n mn hc ti xin gi li cm n n:
Tin S L Th Thy Tin B mn Cng ngh Sinh Hc, i hc Bch
Khoa Tp HCM hng dn tn tnh cng nh cho nhng li khuyn b
ch ti c th hon thnh tt n.
Cc thy c trong b mn Cng Ngh Sinh Hc truyn t cho ti nhng
kin thc hu ch gip ti c c nhng nh hng r rng trong khi thc
hin n.
Cc bn sinh vin lp HC07BSH cng hc tp, trao i kinh nghim v
gip ti.
Ti xin chn thnh cm n mi ngi.

MC LC
MC LC ....................................................................................................................... i
CC T VIT TT......................................................................................................iii
DANH MC HNH....................................................................................................... iv
DANH MC NH ........................................................................................................ vi
DANH MC BNG .................................................................................................... vii
CHNG 1: M U.................................................................................................. 1
CHNG 2: TNG QUAN .......................................................................................... 3
2.1

Tng quan v cy Thng .............................................................................. 3

2.1.1

Gii thiu chung v h Taxaceae .................................................................. 3

2.1.2

Gii thiu chung v chi Taxus ....................................................................... 9

2.1.3

Mt s loi Thng trn th gii ............................................................. 16

2.1.3.1 Taxus baccata ........................................................................................... 16

2.1.3.2 Taxus brevifolia ........................................................................................ 18
2.1.3.3 Taxus canadensis...................................................................................... 19
2.1.3.4 Taxus cuspidata ........................................................................................ 21
2.1.3.5 Taxus floridana ........................................................................................ 23
2.1.3.6 Taxus globosa ........................................................................................... 25
2.1.3.7 Taxus chinensis ........................................................................................ 26
2.1.3.8 Taxus wallichiana .................................................................................... 28
2.1.4

Tnh trng hin ti v bo tn loi Thng Vit Nam.......................... 29

2.1.4.1 Thng Bc (Taxus chinensis Pilg.) ..................................................... 30

2.1.4.2 Thng Nam (Taxus wallichiana Zucc.).............................................. 33
2.2

Taxol .................................................................................................................. 36

2.2.1

Lch s nghin cu ...................................................................................... 36

2.2.2

Cu to Taxol ............................................................................................... 39

2.2.3

Con ng sinh tng hp Taxol .................................................................. 40

i

2.2.4

C ch tc ng ca Taxol .......................................................................... 45

2.2.5

Mt s phng php tng hp hp cht Taxol ........................................... 49

2.2.6

ng dng hp cht Taxol trong iu tr bnh ung th ................................ 57

Chng 3: MT S KT QU VI NHN GING TAXUS ................................ 64

3.1

Vi nhn ging Taxus baccata L. ...................................................................... 64

3.1.1

Mi trng nui cy .................................................................................... 64

3.1.2

Cc giai on ca qu trnh vi nhn ging .................................................. 65

3.1.3

Kt lun ........................................................................................................ 70

3.2

Vi nhn ging Taxus wallichiana Zucc. .......................................................... 70

3.2.1

Mi trng nui cy .................................................................................... 71

3.2.2

Cc giai on ca qu trnh vi nhn ging .................................................. 73

3.2.3

Kt lun ........................................................................................................ 79

Chng 4: KT LUN ................................................................................................ 80

TI LIU THAM KHO ............................................................................................ 81

ii

CC T VIT TT
FDA
U.S. Food and Drug Administration Cc qun l Thc Phm v Dc
Phm Hoa K.
CITES

Convention on International Trade in Endangered Species Cng c

thng mi quc t i vi cc loi gp nguy him.

IUCN

International Union for Conservation of Nature T chc bo tn thin

nhin th gii.
NCI

U.S National Cancer Institute Vin ung th quc gia Hoa K.

CRADA

Cooperative Research and Development Agreement Hip nh hp tc

nghin cu v pht trin.

BMS

Bristol-Myers Squibb.

GGPP

Geranylgeranyl pyrophosphate.

IPP

Isopenthenyl diphosphate.

DMAPP

Dimethylallyl diphosphate.

10-DAB

10-deacetylbaccatin III.

DAB

Baccatin III.

NAA

-naphthaleneacetic acid.

IAA

Indole-3-acetic acid.

IBA

Indole-3-butyric acid.

BA

6-benzyladenine.

iii

DANH MC HNH
Hnh 2.1: Amentotataxus argotaenia Pilger. ...................................................................... 5
Hnh 2.2: Austrotaxus spicata Compton. ............................................................................ 6
Hnh 2.3: Torreya nucifera (L.) Siebold & Zucc. ............................................................... 6
Hnh 2.4: Pseudotaxus chjenii (Cheng) Cheng. ................................................................. 7
Hnh 2.5: Taxus cuspidata Siebold & Zucc. ....................................................................... 7
Hnh 2.6: Phm vi phn b ca h Taxaceae. .................................................................... 8
Hnh 2.7: Phm vi phn b ca mt s loi thuc chi Taxus. .......................................... 11
Hnh 2.9: Phm vi phn b ca Taxus baccata khu vc Bc M.................................. 17
Hnh 2.11: Phm vi phn b ca Taxus brevifolia khu vc Bc M. ........................... 19
Hnh 2.13: Phm vi phn b ca Taxus canadensis khu vc Bc M. ......................... 21
Hnh 2.15: Phm vi phn b ca Taxus cuspidata khu vc Bc M. ........................... 23
Hnh 2.17: Phm vi phn b ca Taxus floridana khu vc Bc M. ............................ 24
Hnh 2.19: Phm vi phn b ca Taxus globosa. ............................................................ 26
Hnh 2.21: Phm vi phn b ca Taxus chinensis. ........................................................... 27
Hnh 2.23 : Thng Bc (Taxus chinensis Pilg.). ......................................................... 30
Hnh 2.24: Phm vi phn b ca Thng Bc (Taxus chinensis Pilg.) Vit Nam. .... 31
Hnh 2.25: Thng Nam (Taxus wallichiana Zucc.). ................................................... 33
Hnh 2.26: Phm vi phn b ca Thng Nam (T. wallichiana Zucc.) Vit Nam. ... 34
Hnh 2.27: Cu trc ca Taxol. ......................................................................................... 39
Hnh 2.28 : Con ng sinh tng hp Taxol. ................................................................... 43
Hnh 2.29: Qu trnh hnh thnh vng oxetane trong sinh tng hp Taxol. ..................... 44
Hnh 2.30: Con ng sinh tng hp Taxol t tin cht gi nh c
polyhydroxyl ha. .............................................................................................................. 45
iv

Hnh 2.31: C ch tc ng ca Taxol i vi t bo gy ung th. ................................. 46

Hnh 2.32: C ch phn ng ca Taxol trong h thng vi ng ca t bo. ...................... 48
Hnh 2.33: Phng php phn tch retrosynthetic ca Nicolaou. ..................................... 50
Hnh 2.34: Bn tng hp Taxol t DAB........................................................................... 57
Hnh 3.4: T l mu cy T. wallichiana trn cc mi trng MS khc nhau. ................. 75
Hnh 3.5: Chiu di chi trung bnh (cm) ca mu cy T. wallichiana trn mi trng
MS sa i khc nhau. ...................................................................................................... 76

DANH MC NH
Hnh 2.8: Taxus baccata L. ............................................................................................... 16
Hnh 2.10: Taxus brevifolia Nutt. ..................................................................................... 18
Hnh 2.12: Taxus canadensis Marsh................................................................................. 20
Hnh 2.14: Taxus cuspidata Siebold & Zucc. ................................................................... 22
Hnh 2.16: Taxus floridana Nutt. ...................................................................................... 24
Hnh 2.18: Taxus globosa. ............................................................................................... 25
Hnh 2.20: Taxus chinensis Rehd. var mariei. .................................................................. 27
Hnh 2.22: Taxus wallichiana Zucc ................................................................................. .29
Hnh 3.1: Vi nhn ging T. baccata.................................................................................. 66
Hnh 3.2: K thut gim cnh c p dng cho Taxus wallichiana. ............................. 71
Hnh 3.3: Thu thp mu cy T. wallichiana. .................................................................... 74
Hnh 3.6: Vi nhn ging in vitro T. wallichiana............................................................... 78

vi

DANH MC BNG
Bng 2.1: Tm tt lch s nghin cu v hp cht Taxol. ................................................ 38
Bng 2.2: Cc nghin cu gn y v kh nng thu nhn Taxol v cc hp cht c
lin quan bng phng php nui cy t bo. ................................................................... 52
Bng 3.1: Thnh phn mi trng dinh dng c s dng cho qu trnh vi nhn
ging T. baccata. ............................................................................................................... 65
Bng 3.2: Nng cc cht iu ha sinh trng v than hot tnh b sung vo mi
trng MS c s dng cho qu trnh tng trng v ko di ca chi.. ................... 72
Bng 3.3: Nng ca cht iu ha sinh trng BA v than hot tnh c b sung
vo mi trng MS cm ng s chi bt nh. ........................................................ 72
Bng 3.4: Nng cht iu ha sinh trng, vitamin B1 v than hot tnh b sung
vo mi trng MS v mi trng MS cm ng s hnh thnh r. ......................... 73
Bng 3.5: T l nhim ph thuc vo loi mu cy. ........................................................ 75
Bng 3.6: nh hng ca chi v thn c ngun gc in vitro v ex vitro trn chiu di
ca chi bn v phn trm mu cy c mang chi. ........................................................... 77

vii

Chng 1: M U

CHNG 1: M U
Cc loi Taxus c bit n t thi c i, gn nh tt c cc b phn ca
n (ngoi tr o ht) u cha cht c i vi con ngi v ng vt.
Trong y hc, Thng l mt loi dc liu qu. T lu trong dn gian, cy
Thng c dng iu tr hen suyn, vin ph qun, nc, mt s bnh
tiu ha, cnh v v dng tr bnh giun a, nc sc ca cy Thng non tr
bnh au u
Nm 1963, cc hp cht c tch chit t v cy Taxus brevifolia, c hiu qu
trong cc nghin cu tin lm sng: chng li cc khi u trong c th. Nm 1971,
Taxol (Paclitaxel) - mt diterpenoid alkaloid, c xc nh l thnh phn chnh
trong hp cht ny (Wall & Wani, 1971) [32]. Taxol thc y c ch lp rp ca
cc tubulin v n nh hiu qu ca vi ng. Hiu qu ca n c kim nh
lm sng iu tr ung th v, bung trng, thc qun, bng quang, ni v c t
cung Taxol c chp thun ln u tin bi Cc qun l Thc phm v Dc
phm Hoa K (U.S. Food v Drug Administration) dng trong iu tr ung th
bung trng vo nm 1992 v ung th v di cn vo nm 1994 (Suffness v Wall,
1995) (theo Croteau, 2006) [16].
Taxol cng c tch chit thnh cng t cc loi khc trong chi Taxus,
cng nh cc b phn khc ca cy bao gm phn hoa, ht ging, l, thn v r
(Vidensek v cng s, 1990; Witherup v cng s, 1990; Fett-Neto v cng s,
1992; Wickremesinhe v Arteca, 1994) (theo D.T. Nhut, 2007) [15].
Vn kh khn nht gp phi trong sn xut Taxol chnh l ngun cung cp
hn ch vi hm lng rt thp. c tnh cn 7000 kg v cy c th sn xut
c 1kg Taxol (Cragg v cng s, 1993) [13]. Mt khc, cy tng trng chm
trong iu kin t nhin, thi gian ng ca ht ging ko di t 1.5 nm n 2 nm
(Steinfeld, 1992) (theo D.T. Nhut, 2007) [15].
Ngoi ra, t cc b phn ca nhiu loi Taxus cn thu nhn c cc hp cht
th cp khc nh lignan, flavonoid, steroid v cc dn sut ca ng. Ngi ta
tm c khong 50 lignan, bao gm neolignan v mt vi terpenolignan. C th
trong T. baccata (European yew), 5 lignan c tm thy: lariciresinol,
taxiresinol, 3-demethylisolariciresinol-9-hydroxyisopropylether, isolariciresinol
1

Chng 1: M U
v 3-demethylisosalariciresinol [27]. Cc nghin cu in vitro ch ra rng
larciresinol v isolarciresinol c hot ng c ch mnh m n nhn t gy hoi
t khi u (tumor necrosis factor- (TNF-)) v taxiresinol c cho l c hiu qu
cao trong bo v chng li cc tn thng d dy.
Hin ti, vic tch chit, kim tra hat tnh sinh hc ca diterpenoid alkaloid
mi cng nh tng hp hay bin i cu trc ca cc taxane c cp trong
nhiu nghin cu. Hng trm cu trc khc nhau ca taxoid c bit n v mt
s c kh nng lm tin cht sn xut Taxol, mt s khc c tc dng tng t
thm ch vt tri hn so vi Taxol. iu ny cho thy tim nng ca nhiu loi
Taxus vn cha c khai thc ht, n ha hn s l ngun cung cp dc liu
quan trng cho iu tr ung th v nhiu loi bnh khc trong hin ti v tng lai.
Vic nghin cu trn cy Thng Vit Nam cn rt khim tn, ch mi
din ra trong khong 10 nm tr li y v tng bc t c mt s thnh cng
bc u ch yu trn lnh vc nhn ging in vitro. Hin nay xc nh c 2
loi Thng l Thng Bc (Taxus chinensis Pilg.), phn b ch yu cc
tnh vng cao nh Sn La, H Giang, Tuyn Quang v Thng Nam (Taxus
wallichiana Zucc.) phn b ri rc mt s tnh Khnh Ha, Lm ng ( Lt,
n Dng) [1]. Tuy nhin, qun th Thng ny trong t nhin rt t, nu dng
chit xut Taxol th s liu thuc cha tr ung th cng ch l mt con s rt
nh (D.T Nhut, 2007). T thc trng ny, cho thy vic nhn ging cy Thng
bng cc phng php in vitro v a ra trng i tr l cn thit cung ng cho
nhu cu iu tr trong nc.
n ny c thc hin vi mc ch a ra mt ci nhn tng quan v cy
Thng trn th gii v Vit Nam, phn loi, c im, phm vi phn b
cng nh v hp cht Taxol, tnh cht l ha, con ng sinh tng hp, c ch v
ng dng ca Taxol trong iu tr ung th. Cui cng, trnh by mt s kt qu
nghin cu nhn ging in vitro trn mt s loi Thng nh Taxus baccata L. v
Taxus wallichiana Zucc.

Chng 2: TNG QUAN

CHNG 2: TNG QUAN

2.1 Tng quan v cy Thng
2.1.1 Gii thiu chung v h Taxaceae
Taxaceae mt h nh chim khong 2% trong ngnh thc vt ht trn, bao gm
5 chi v 25 loi c phn b ch yu Bc bn cu [16].
2.1.1.1 Tn thng gp
Yew (Hils, 1993) [26], Taxaces [French], Eibengewchse [German], taxceas
[Spanish], [Russian], hong dou shan ke [Chinese],
[Japanese] [31].
2.1.1.2 M t c im chung
Cy thng xanh, n tnh khc gc (i khi n tnh cng gc nh loi
Torreya v Taxus canadensis), cy thn g hoc cy bi, v mng v d bong hoc
v th, dy v b cht, mu nu hoc nu m, i khi xut hin tinh th calci
oxalate kt tinh, g khng c nha, vi cu trc xon c trn vch th cp ca
trc t bo dng ng (thng khng c trong Austrotaxus), cnh lan rng n
thng ng v tng dn chiu cao, cnh i xng hoc xen k, khng c lng, mu
nu hoc mu xanh l, mang l gn nh khng c cung l [16].
L tn ti t 2, 3 hoc 4 nm, thng thng l 2 nm, thng c nh, ni bt
vi decurrent di, xen k (ngoi tr Amentotaxus), thng ti thng mi mc, mng,
nhn mi hoc nhn u, khng c lng, i khi c nh trn gn gia hoc mp l,
cung l di 1 - 2 mm hoc khng c cung l, mp l lin, cun ngoi (revolute),
mt trn l mu xanh m, mt di mu xanh m, xn vng hoc vng xanh, c
kh khng, dc theo 2 bn, mu lc lam hoc mu xanh nht ti mu hung, l n ,
gn gia ni, to thnh rnh hoc khng r rng mt trn, hin r hn mt
di, nha c trong cc ng n (Amentotaxus), nm ri rc trong nhu m hoc
khng c [16].
Nn c c cui l hoc nch l, n l hoc thnh tng cm t 2 4 nn,
khng cung hoc c cung, hnh cu hoc hnh tr thun di khi cy cn non,
hnh tr hoc hnh nn khi cy trng thnh, di 2 - 10 mm, vy khng sinh sn (
8) di cc l bo t nh (vi 2 vy khng sinh sn xen gia cc l bo t nh
3

Chng 2: TNG QUAN

trong Pseudotaxus), l bo t nh t 3 - 14 (rt nhiu trong Torreya), ti bo t
thp v i xng ta trn (trong Taxus Pseudotaxus, v Austrotaxus) hoc ti bo
t cao (ch c ti bo t dc theo trc pht trin y ) v sp xp mt lng
bng (trong Amentotaxus v Torreya), di 1 - 3 mm, c cung (tr Austrotaxus),
hnh khin hoc c nh dy, ti bo t sn xut giao t c t (2 -) 3 - 9, phn hoa
khng c bng hoc cnh (ngoi tr trong Amentotaxus argotaenia), c cung
hoc khng c cung, ht (granular) mng hoc sn si, mng ngoi ca bo t
trong 2 lp c dy bng nhau [16].
Nn ci bao gm mt nn nm n c tn cng, khng cung hoc c
cung di khong 1.5 cm, vy khng c kh nng sinh sn, 6 - 14, vng xon sp
xp i xng hoc dng ch thp, non trng thnh hay ht c bao quanh bi
v o (mt phn tht ca cc l bo t ln (c quan sinh sn ci) c pht trin t
trc hoc nh ca nch chi mang non), v o c dng hnh cu, hnh trng hoc
thun di, 5 - 30

10 - 15mm, c mi thm, mm, c mu , trng, xanh hoc

tm, ht kh v nutlike (ging ht), vi 2 l mm [16].

S lng nhim sc th, n = 12, c trng hp n = 11 (trong Torreya
californica v T. nucifera, c th c Amentotaxus formosana), hoc trng hp
him gp n = 7 ( A. argotaenia) hoc khng m c (Austrotaxus,
Pseudotaxus) [16].
2.1.1.3 Phn loi:
H Taxaceae bao gm 5 chi v 25 loi (bao gm c 2 loi lai nhn to)[16].

Chng 2: TNG QUAN

Kha phn loi cc chi trong h Taxaceae [16]:
1. Nn c to thnh chm, l rng 5 - 9 mm.
2. L i xng, nn c nm tn cng hoc gn tn cng, qun bo xon
dy . ....................................................................................... Amentotaxus
2. L sp xp hnh xon c, nn c nch l, t bo ng khng xon
dy
................................................................................................ Austrotaxus
1. Nn c nm n c, l rng < 5mm.
3. o ht ng li nh, che ch cc ht ging, cnh i xng, gn
gia khng ni mt trn ca l ................................................. Torreya
3. o ht m nh, l ra cc ht bn trong, gn gia ni ln b mt
trn ca l.
4. o ht mu trng, nn c c vy khng sinh sn, nh ch c
mp l
....................................................................................... Pseudotaxus
4. o ht mu , nn c khng c vy khng sinh sn, nh dc theo
trc gn l ................................................................................. Taxus

Hnh 2.1: Amentotataxus argotaenia Pilger [16].

A. Cnh, nn ci trng thnh rng,v nn ci cha trng thnh cn li.
B. Nn c. C. Nn ci v o ht. D. B mt l (tri) v (phi) dc theo trc.
5

Chng 2: TNG QUAN

Hnh 2.2: Austrotaxus spicata Compton [16].

A. Cnh vi nn ci v o ht. B. Cnh vi nn c. C. Nn c.
D. B mt l (tri) v (phi) dc theo trc.

Hnh 2.3: Torreya nucifera (L.) Siebold & Zucc [16].

A. Cnh mang nn c. B. Nn c. C. Cnh mang nn ci cha trng thnh,
trng thnh v o ht . D. B mt l (tri) v (phi) dc theo trc.

Chng 2: TNG QUAN

Hnh 2.4: Pseudotaxus chJenii (Cheng) Cheng [16].

A. Cnh. B. Nn c. C. Nn ci v o ht.
D. B mt l (tri) v (phi) dc theo trc.

Hnh 2.5: Taxus cuspidata Siebold & Zucc [16].

A. Cnh mang nn c. B. Nn c khng m.
C. Cnh mang nn ci. D. Nn ci v o ht.
E. B mt l (tri) v (phi) dc theo trc.
7

Chng 2: TNG QUAN

2.1.1.4 Phn b :
H Taxaceae c phn b Bc bn cu. Chu u: t Anh ti Bc Iran. Chu
: Nga, Hn Quc, Nht Bn, Trung Quc, i Loan, Himal, n , Myanmar,
Vit Nam, Philippines. New Caledonia. Khu vc Bc M: ng Nam Arkansas ti
California, ng Canada, ng M, Florida, Mexico, Guatemala, El Salvador.
Khu vc Nam M [31].
Hu ht cc chi trong h Taxaceae rt him hoc phn b hn ch mt s
vng nht nh, tuy nhin c mt s loi thuc chi Taxus phong ph v phm vi
phn b rng hn.
Torreya taxifolia Am. v Taxus florina Nutt. thuc nhm thc vt c nguy c
tuyt chng cao phn b hn ch Florida v Georgia (Godfrey & Kurz, 1962;
Ward, 1979) (theo Cope, 1998) [16]. Taxus globosa Schldl. l mt loi c hu qu
him ca min Trung Mexico. Austrotaxus v Pseudotaxus ch c tm thy
New Caledonia v tnh Chit Giang (Trung Quc), tng ng vi tng loi.
Torreya californica ch trng c min Bc California v min Nam Oregon.
Ba loi khc thuc chi Torreya ch phn b hn ch mt s tnh ca Trung Quc,
v Torreya nucifera ch yu c trng Nht Bn v Hn Quc [16].
Ch c Torreya v Taxus c trng theo quy m ln M. Tuy nhin,
Torreya phn ln c trng trong cc vn thc vt v gn nh tt c chi Taxus
c trng lm cnh nh T. baccata, T. cuspidata, v dng lai gia chng, T.
media, c trng rng ri trong vng ng Bc v Trung Ty M.

Hnh 2.6: Phm vi phn b ca h Taxaceae [16].

8

Chng 2: TNG QUAN

2.1.2 Gii thiu chung v chi Taxus:
Chi Taxus (Taxaceae) t lu c ch nh bao gm 7 - 12 loi hoc phn
loi (Cope, 1998; Farjon, 1998, 2001; Pilger 1903, 1916; Silba 1984, 1986; Spjut
1992), tuy nhin, Spjut (2000c; www.worldbotanical.com) ngh cng nhn
chi Taxus bao gm 24 loi, 54 ging [26].
H thng phn loi ca Taxus c tranh lun t rt lu do qun th loi ny
khng c cch ly v mt sinh sn m ch c cch ly a l (Farjon 1998; Pilger
1903; Silba 1984). Cc chi phn b t vng n i Bc M ti cn nhit i Trung
M, t vng n i - u ti vng cn nhit ti ng Nam (Cope, 1998) [16].
2.1.2.1 Tn thng gp
Yew.
2.1.2.2 M t c im chung
Taxus spp. l cy thng xanh, n tnh cng gc hoc n tnh khc gc. V
mu nu , c vy. Nhnh thng sau r xung, mu xanh l cy hoc mu
vng lc khi cn nh, sau chuyn thnh mu nu khi trng thnh.
L thng xut hin 2 hng, mm do, kh khng xp dc theo trc, gia 2
hng c cc vng nht mu hn, nh l mm, nhn, khng sc cnh, c nha
trong cc ng n [31].
Nn phn hnh cu, mu vng, vi 4 - 16 l bo t hnh khin, mi ci mang 2 9 ti bo t. C mt non. Ht cng trong ma u tin, mu nu; o ht mu
cam hoc ti, mm, nhy, dy, o ht m nh l ra cc ht cng bn
trong. Phi nh ng nht. n = 12. (Hils, 1993) [24].
2.1.2.3 c im sinh trng:
Taxus spp. l loi a m, pht trin ch yu di tn rng, mi trng mt m,
kh hu n i hoc cn nhit i, cao trn mc nc bin (rng n i) n
khong 3000 m (rng cn nhit i). Hnh thc sinh trng c th l cy thn g
hoc cy bi. Cy c nhiu cnh v tn rng c th hp thu c ti a nh sng
di tn rng. Cy c th chuyn thnh dng cy bi nu chi ngn b tn thng
v c thay th bng cc chi bn hoc mc chi mi [31].

Chng 2: TNG QUAN

Trong nhiu ti liu cng b cho thy Taxus spp. c thi gian sinh trng
chm v c tui th cao. c th t c kch thc trung bnh ca loi, thi
gian sinh trng c th ln n hn 100 nm. Ht nm ring l, tc pht tn ht
ging chm v sc sng ca ht ging rt thp.
Cy con thng chm pht trin trong iu kin t nhin cng nh trong trng
trt. Qu trnh nhn ging cng rt chm (thng thng hai nm l thi gian cn
thit cho s ny mm t nhin ca ht ging, t 2 n 3 thng nu nhn ging
nhn to bng phng php gim cnh).
Chiu cao ca mt s loi Taxus nh T. wallichiana (Himalayan yew) t 20 30 m ng knh 1 (1.5) m (Nguyn v cng s, 2004). T. brevifolia (Pacific yew)
cao khong 20 m (Farrar, 1995). T. baccata (English yew) chiu cao t c t
12 ti 25 m (Krussman, 1983). T. floridana (Florida yew) khi trng thnh chiu
cao ti a khong 5 m (Vance v Rudolf, 1974). T. canadensis (Canada yew) l
cy bi vi tn rng, chiu cao him khi vt qu 2 m (Farrar, 1995) [26].
Tui th ca loi Taxus spp. cao, thng thng t 250 - 500 nm tui. Loi T.
baccata c tui th trong bnh khong 250 nm, tuy nhin vn c nhng trng
hp ring bit tui th ca loi ln ti 1000 nm tui (Vance v Rudolf, 1974)
(theo Spjut, 2007) [26].
2.1.2.4 Phn loi v phm vi phn b:
y trnh by mt kha phn loi gm 24 loi v 26 ging ca Taxus da
trn bn cht ha hc (sn phm ph) v s khc bit hnh thi. M t v mu vt
c trch dn c cung cp ch yu trn d liu a l sinh hc ca Taxus (Spjut
2007). N cng bao gm cc nghin cu hin nay trn cc ging T. brevifolia, T.
caespitosa Nakai, T. canadensis, v T. umbraculifera, khng c cp trong d
liu ca Spjut (2007) [26].
Taxus c phn loi thnh 3 nhm, trong nhm bao gm 2 phn nhm v 2
lin minh (alliance) da vo hnh dng t bo biu b l, s pht trin ca nh, v
mu sc ca cc dy kh khng vi cc vng ln cn ch cha t bo biu b (Spjut
1998b). Nhm Wallichiana bao gm phn nhm Wallichiana v Chinensis, nhm
Baccata bao gm 2 lin minh Baccata v Cuspidata, nhm Sumatrana [26].
Lin minh da trn cc n v phn lai c chung cc c im hnh thi trong
khu vc a l, trong khi phn nhm v nhm khng c cch ly v mt a l.
10

Chng 2: TNG QUAN

Trong cc loi lin minh hoc nhm, loi lin hp (complex) c cng nhn
phn bit, v d, s phn b i xng ta trn ca l (T. umbraculifera complex)
so vi l xp thnh 2 hng (T. cuspidata complex).

Hnh 2.7: Phm vi phn b ca mt s loi thuc chi Taxus [16].

Cha kha phn loi loi v ging ca chi Taxus [26].
NHM WALLICHIANA.
1. PHN NHM 1A WALLICHIANA.
1. S lng kh khng : 12 kh khng/dy; ng Himalaya, Ty Nam Trung
Quc.
2. Cm chi n nh, tng i ln, di 2 - 3 mm, l phng vi gn l ring
bit; Myanmar. ......................................................................... T. suffnessii
2. Cm chi n nh khng c gn l hoc gn nh khng c, lm xung,
chiu di > 2 mm v rng 1 mm; ng Himalayan, Nepal ti Ty Nam
Trung Quc (var. yunnanensis c phn bit bi dc theo mp l c cc t
bo di v hp, gn mp ngn hn, t bo nh rng hn vi nh trung gian,
hoc l c hnh mi mc, Ty Nam Trung Quc). ............... T. wallichiana
1. S lng kh khng: < 12 kh khng / dy.
3. T bo biu b dc theo trc hp v ngn ti dy kh khng mp l, phn
ln c cng kch thc dng ch T, hnh thoi v ln sng (ln theo vch
t bo) trn b mt dc theo trc, i khi l hnh ch nht. .... Taxus globosa
4. Nh ni trn hu ht b mt l dc theo trc, cc t bo mp ch yu
dng ln sng; t El Salvador ti Ty Bc Mexico.
..............................................................................T. globosa var. globosa
11

Chng 2: TNG QUAN

4. Nh t ni hn trn gn gia v mp l trn dy kh khng, t bo
mp l hnh ch nht; Mexico (Nuevo Leon/Tamaulipas, Veracruz),
U.S. (Florida).
.......................................................................... T. globosa var. floridana
3. T bo biu b ngn t ngt v rng hn mp l, thng cao hn chiu
rng dng ch T.
5. Kh khng 7 - 10( - 12) hng; Trung Quc (Vn Nam, T Xuyn).
..........................................................................................................T. florinii
5. Kh khng 4 - 7( - 9) hng; ng Bc M. ........................ T. brevifolia
6. Nn thun di, thng gn trn cung hp.
................................................................ T. brevifolia var. polychaeta
6. Nn khng c cung.
7. Cy thn g, chi mc ngu nhin. .... T. brevifolia var. brevifolia
7. Cy bi, thng mc thnh tng cm.
.............................................................T. brevifolia var. reptaneta
1. PHN NHM 1B CHINENSIS.
1. B mt l kh, gn nh cng mu, hoc mu vng lc dc theo trc.
2. Cm chi d thy ti gc ca cnh; Vn Nam, T Xuyn.
................................................................................................. Taxus sp. SCU
2. Cm chi nh, ch c vt hoc khng c cnh ca cy 1 nm tui.
3. L dng trn, thng l thun; Vit Nam, Trung Quc. .....T. chinensis
3. L nhn; Vit Nam, Philippines, Trung Quc (i Loan), Indonesia.
........................................................................................... Taxus sp. REH
1. B mt l kh, khng mu hoc mu vng cam dc theo trc.
4. L dng trn, b mt nhn; Vn Nam, T Xuyn. ..............Taxus sp. OCR
4. L nhn, mn, bng trn b mt khi l kh.
5. L phn b song song gn gc phi; Nepal, Ty Bc n ,
Malaysia, Trung Quc (i Loan, Vn Nam), Philippines.
.............................................................................................T. phytonii
5. L phn b theo gc xin, an cho trong cc mu p; Myanmar,
China (Phc Kin, i Loan), Philippines, Indonesia (Sulawesi,
Sumatera).
............................................................................................. T. obscura
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NHM SUMATRANA
1. L kh mu cam bc, gn nh thng bc mu tng phn vi mu xanh
m hoc mu ng b mt l, xon xin vi trc thn, thng un ngc
dc theo phin l ti nh nhn, vi gn gia trn u trn c 2 b mt; Ty
Bc n ti Trung Quc (bao gm i Loan). ............................ T. kingstonii
1. L kh mu xanh , hoc mu mu phai dc theo mp l v trn gn gia,
xon gn nh vung gc vi trc, cc mp l lin k thng song song, l trn
ti nhn, gn gia ni cao v ngn, hoc phng, thng ch c mt gn, mng
v mm; Trung Quc.
2. L thun di, trn ti nhn hoc hnh ellipse nhn, t bo biu b ln
(mammillose) dc theo trc ln hn gn gia, c hnh thang ngn; Nam
Trung Quc. .................................................................................... T. mairei
3. Cnh mm, phn thnh nhiu nhnh, l st nhau, l song song dc theo
mt bn ca cnh, ht mu ta. .................................T. mairei var. mairei
3. Cnh cng, i khi c phn nhnh, l cch qung khng u dc theo
cnh, ht mu nu. .................................................T. mairei var. speciosa
2. L di thng hnh mi mc, nhn, cc t bo biu b dng ch T (T-sect)
khng ln hn cc t bo dc theo gn gia, hnh ch nht; t ng
Himalaya ti Indonesia, Philippines.
4. L kh phng hoc li trn b mt dc theo trc. ................... T. celebica
4. L kh nhn. ....................................................................... T. sumatrana
NHM BACCATA
1. Trn b mt hng trc, thng c nhiu nh gia mp l v dy kh khng, c
khong 8 hng t bo mp l, nh trung gian, thnh t bo mng, l thng cong
xung dc theo phin l (ngoi tr T. contorta var. mucronata); a Trung Hi,
Ty Himalaya.
.................................................................................................... T. baccata Alliance
2. Tht l vi cc d bo (cc t bo nhu m vi thnh t bo mu ), lin kt
lng lo; t Ty Bc ti min Trung Himalayas. ......................... T. contorta
2. T bo nhu m thng c mu ti, khng c mu , lin kt cht.
2a. Cy hnh tr, l ta trn, un ngc li. .................................. T. fastigiata
2b. Thc vt thay i i theo mi trng sng (thn g hoc cy bi ty
theo mi trng sinh trng), l chng cho, khng phn b ta trn.
2b. B mt l bc mu, phin l un ngc, li, cun ngoi. ..... T. recurvata
13

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2b. L cng mu, thng hoc hnh lim, phng hoc cun ngoi. . T. baccata
2c. L phn b pha ngoi dc theo hai bn ca cnh trn mt mt phng,
xp thnh hai dy.
2d. L khng xp chng cho, cnh phn thnh nhiu nhnh.
..............................................................................T. baccata var. baccata
2d. L chng cho, cnh khng theo quy lut.
.................................................................... T. baccata var. elegantissima
2c. L mc thng hoc ta trn vo phn di ca cnh.
2e. Cnh v l c mu cam vng khng u, cnh thng un ngc, l
dng ng u cnh. .......................................... T. baccata var. glauca
2e. Cnh c mu xanh l, mu vng xanh hoc mu xanh m ng
nht, cnh khng un ngc, lan rng hoc r xung, l thay i.
2f. Cnh ch yu c hnh lng chim. .............. T. baccata var. variegata
2f. Cnh nhiu hn ngn, xut hin s pht sinh phn cnh hoc
chia nhnh.
2g. L vng xanh, rng vo nm th 3, cnh phn thnh nhiu nhnh.
................................................................. T. baccata var. pyramidalis
2g. L xanh m, lin tc, cnh phn nhnh hnh lng chim hoc
hnh chp. ............................................ T. baccata var. dovastoniana
2. L nhn gia dy kh khng v mp l, thng khng c nh qua t 8-24 hng
t bo, cc nh thng sinh trng mt trn ca t bo gn thnh t bo,
phin l cong ln trn hoc un t ngt hoc cong xung di dc theo cung
l; ng , ng Bc M, a Trung Hi. ................... T. cuspidata Alliance
3. L tri rng nh cung l hp, cung l cong t ngt, kh khng (4- ) 5 9(- 11) hng trn mt dy, nh l thng khng r, cy thp, cy bi tng
trng chm. ..............................................................T. canadensis complex
4. L nhn, thon dn cho ti mp l ging dng ch T, thng cun ngoi
dc theo mp l, b mt dc trc vi gn gia lt p v t bo biu b
hnh ch nht; ph bin ng Bc M, xut hin t hn a Trung
Hi, Marocco, B o Nha (Madeira), Ty Ban Nha, Thy in,
Estonia.
.................................................................. T. canadensis var. canadensis
4. L trn ti nhn, mp trn trong dng ch T, khng cun ngoi, vi gn
gia bng v t bo biu b hnh thang hoc hnh ch nht.
5. L nhiu, dng ng, un ngc li; ng Bc Amer, Madeira.
..................................................................... T. canadensis var. minor
14

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5. L tha, thng, gn nh vung gc; Anh, Madeira, Php, Thy
in, Na Uy, Slovenia, M (him). ....... T. canadensis var. adpressa
3. Cung l cong t, un cong gn ch giao vi phin l, phin l thng p vo
cnh nh, kh khng (7-) 9 - 14(- 17) hng trn 1 dy, nh thng ni bt trn
dy kh khng, cy thay i theo mi trng sng.
6. L xp thnh hai hng ti u cnh, ht c gc cnh. . T. cuspidata complex
7. Cnh ngn v chia nhiu on, l trn cnh nh, cng hn khi kh, ch
yu thuc nhm cy bi; ng Bc Chu . .................... T. biternata
7. Cnh ch yu hnh lng chim di, l dng ng, cun ngoi ng u
dc theo mp l khi sy kh, cy bi hoc cy thn g-xut hin khi
thch nghi tt vi mi trng sng; Nht Bn, Hn Quc.
............................................................................................... T. cuspidata
6. L hai hoc nhiu hng xp chp hnh ch thp hoc ta trn, ht trn.
............................................................................. T. umbraculifera complex
8. L dng ng v xp gi ln nhau. ...................................... T. caespitosa
9a. Cnh dn thng ng vi nhiu cnh nh v ngn, l ch yu ta
trn; Nht Bn. ........................................ T. caespitosa var. caespitosa
9a. Cnh phn b rng, cnh nh vi s ln ln hai hng v dng ng
mt pha.
9b. Cnh lan rng pha ngn , l thun hoc thng, rng 2 - 4 mm;
Hn Quc, Nht Bn, ng Nam Nga, ng Bc Trung Quc.
...................................................................... T. caespitosa var. latifolia
9b. Cnh c ph vi cnh nh dng ng, l thng, tng i hp,
rng 2 mm; Hn Quc, Nht Bn. ...... T. caespitosa var. angustifolia
8. L xp cho hnh ch thp v un ngc..................... T. umbraculifera
10a. Cy bi phng, xp lp.
.......................................................... T. umbraculifera var. microcarpa.
10a. Cy bi dng chm cu, dng ct hoc cy thn g.
10b. Cy bi thp, trn. .............................. T. umbraculifera var. nana
10b. Cy bi cao hoc cy thn g.
10c. Dng ct vi cnh nh dng ng. T. umbraculifera var. hicksii
10c. Dng kim t thp, cnh sp xp ln xn.
.................................................. T. umbraculifera var. umbraculifera

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2.1.3 Mt s loi Thng trn th gii
2.1.3.1 Taxus baccata
a. Tn thng gp
European, English, hoc common yew (Hartzell, 1991) [31].
b. M t c im [16, 26]
Taxus baccata ch yu l cy n tnh khc gc, cy cao t 10 20 (- 40) m v
ng knh ln n 4 m. Tn cy c hnh kim t thp, thay i ty theo tui
cng nh iu kin mi trng. V mng, c vy, mu nu. L phng, sp xp 2
hng theo hnh xon c. 10 - 40

2 - 3 mm, c mu xanh m.

Nn c hnh cu, ng knh t 3 - 6 mm, phn hoa rng vo u ma xun.

Ht ging hnh nn bao gm mt ht c bao xung quanh bi o ht c mu
ti, di t 8 - 15 mm.
Khong t 1 - 2 chi c 1 o ht, 10
-7

6 mm, mu ti, ht c hnh ellip, 6

5 mm, mu xanh nu hoc mu nu. o ht trng thnh sau 6 - 9 thng k

t khi th phn, v ht c pht tn ch yu nh chim.

Hnh 2.8: Taxus baccata L [32].

A. Bi cy. B. Nn c. C.o ht v ht ging. D.Ht ging. E. V v thn cy.
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Chng 2: TNG QUAN

c. Phm vi phn b
T Anh cho ti pha Bc Iran (Silba, 1986); c trng rng ri khu vc Bc
M (Hils, 1993) [24]. Ngoi ra cn phn b ri rc Croatia, Slovenia v Balan
[31].

Ghi ch:
Taxus baccata

Hnh 2.9: Phm vi phn b ca Taxus baccata khu vc Bc M [32].

d. Cng dng [31]
T xa xa, g c dng lm cung tn s dng trong c sn bn cng nh
tronng chin tranh cho ti khi sng c pht minh. Taxus baccata cng c s
dng lm cht c tm vo u mi tn gy c cho qun i phng cng nh
cc loi ng vt. Trong i sng hng ngy, g cy c s dng lm ct bum,
trc bnh xe, n, lc
Trong y hc, T. baccata c s dng iu tr vt rn cn, hi chng s
nc (hydrophobia) v mt s bnh lin quan ti tim mch. Hin nay, Taxus
baccata ch yu c s dng lm cy cnh.
Cc nghin cu chng minh, Taxus baccata c cha hp cht Taxol, mt
hp cht dng trong iu tr ung th. L v ht c cha nhiu alkaloid, c bit l
taxine, cc c, d b thy phn thnh hydrotaxine, ngoi ra cng cha cc hp
cht glucoside v taxicatine. L gi v kh c hn so vi l cn non, thng
xuyn gy c cho ng vt, nga v la c th b cht trong vng mt gi sau khi
n. i vi ngi, n tc ng ln h tiu ha, h hp, tim mch v h thn kinh
c th dn ti t vong. Triu chng bao gm: nhp tim v nhp th tng nhanh, tip
n tim p chm, h huyt p, au bng, au d dy, tiu chy cp, chng mt, co
git, hn m v t vong.
17

Chng 2: TNG QUAN

o ht mu c th s dng lm thc n, n c v ngt v nhy, c tc dng
li tiu, nhun trng.
2.1.3.2 Taxus brevifolia
a. Tn thng gp
Pacific hoc western yew, mountain mahogany (g g ni) (Peattie, 1950) [31].
b. M t c im [26]
Taxus brevifolia l cy n tnh khc gc, cy bi hoc cy g nh, chiu cao
t 15(- 25) m, ng knh 50(- 140) cm. Thn cy thng, tn c hnh nn. V cy
c vy, vy bn ngoi mu nu tm hoc nu ta, vy bn trong mu hoc
tm, vy c kch thc khc nhau (20 - 50 mm). Nm u tin, chi non c mu
xanh c bao ph bi l vy, cnh cy c mu nu ging v cy vo nm th
3. L xanh, thng, nhn, di 8 - 35 mm, rng 1 - 3 mm, mm do, c hnh dng
ging ci lim vi sng l hp mt trn ca l, c t 5 - 8 dy kh khng, c nh.
Cung l di 5 - 8 mm. Nn c nm n c hoc thnh cm, cn non c mu
xanh, hnh cu, ng knh 1,5 mm. Ht hnh cu, 5 - 6.5 mm, trng thnh vo
cui ma h, c bao trong cc o ht mu , ng knh 10 mm. G cng.
(Hils, 1993; Peattie, 1950).

Hnh 2.10: Taxus brevifolia Nutt [32].

A. Cnh. B. Nn c cha trng thnh. C. o ht. D.Ht ging. E. V v thn cy.
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c. Phm vi phn b
M: ng Nam Alaska, Montana, Idaho, Oregon, Washington v Bc
California; Canada: Columbia, Alberta (dy Rockie) [31].
c tm thy cao 0 - 2200 m c m cao nh ven sui, sn ni, hm
ni v cc vnh nh (Hils, 1993) [24]. Trong rng l kim, cy ch yu pht trin
nh cy bi di tn rng. Tuy nhin trong rng l rng, cy tng v kch thc
cng nh chiu cao, thay i hnh thi sinh trng thnh cy thn g.

Ghi ch:
Taxus brevifolia

Hnh 2.11: Phm vi phn b ca Taxus brevifolia khu vc Bc M [32].

d. Cng dng [31]
Dn tc bn a vng Ty Bc Thi Bnh Dng coi loi thng ny l loi cy
rt c gi tr, c s dng lm v kh nh cung, lao mc ( nh c), lm mi
cho, cc dng c sinh hot hng ngy nh mung, a, kim Dng trong iu tr
vt thng, h nhai l v p ln vt thng. Cy c lt v, sy kh v c
pha vi nc si lm thuc tr bnh phi.
V cy ca Taxus brevifolia l mt ngun cung cp Taxol t nhin, dng trong
iu tr nhiu loi ung th (c bit l ung th bung trng) [22].
2.1.3.3 Taxus canadensis
a. Tn thng gp
Canadian yew, American yew, cy c cn (ground-hemlock), li du Canada,
sapin trainard (Hils, 1993) [24].
19

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b. M t c im [26, 31]
Cy bi, cao khong 2 m, thng l cy n tnh cng gc, thp, cnh rm rp,
lan rng. V c mu , rt mng. L 1 (2.5) cm

1(2.4) mm, nhng vng

khng c nh dc theo cc dy kh khng l c mu xanh nht, dy kh khng c

mu xanh m hay hoc vng xanh. Mt trn l c mu xanh sng, mt di l c
mu vng xanh, i khi tr thnh mu hoc tm trong ma ng. Cung l di
0.5 - 1 mm. nh l trn nhiu hn nhn. Gn gia khng r rng mt trn cu
l.
o ht hnh cu hoc gn nh hnh cu, 6(8) 6 mm, mu cam hoc ti.
Ht c hnh trng, 4 - 5 mm, c gc cnh nh. 2n = 24. Ht trng thnh vo
cui h u thu (Hils, 1993) [24].
Ht pht tn ch yu nh cc loi chim.

Hnh 2.12: Taxus canadensis Marsh [32].

A. Cnh. B. Nn c. C. o ht trng thnh v cha trng thnh. D.Ht ging.
E. V v thn cy.

20

Chng 2: TNG QUAN

c. Phm vi phn b
Taxus canadensis l loi bn a ca min ng Hoa K v Canada, pha nam
ti Virginia, pha Bc ti Manitoba v Iowa.
Thng c tm thy cao t 0 1500 m, di tn rng (rng l kim,
rng l rng hoc rng hn hp), hoc trong m ly, khe ni v cc dy ni .

Ghi ch:
Taxus canadensis

Hnh 2.13: Phm vi phn b ca Taxus canadensis khu vc Bc M [32].

d. Cng dng [31]
Tt c cc b phn ca cy (ngoi tr o ht) c c tnh nh, cy c s dng
rng ri bi ngi dn bn x. H s dng cc b phn ca cy nh l cnh lm
thuc tr bnh thp khp, vim khp, mt s bnh lin quan ti ng tiu ha, v
s dng lm dc liu xng iu tr cm st.
Taxus canadensis c chng minh l mt ngun cung cp hp cht Taxol
cng nh c cha 2 loi taxane c th s dng tng hp Taxol. Vi mt hm
lng cao hp cht taxane trong Taxus canadensis, n tr thnh mt trong nhng
ngun cung cp hp cht taxane c gi tr nht trong ngnh cng nghip dc
phm.
2.1.3.4 Taxus cuspidata
a. Tn thng gp
Japanese yew, Ichii, Onko (Japanese) (Hartzell, 1991) [31].
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b. M t c im
Cy cao khong 20 m, tn cy hnh kim t thp, trong canh tc ch yu l cy
bi. V cy c vy, mu nu hoc nu sm. Cnh mu nu. Chi nm u
tin c mu vng xanh, ngn chi hnh trng thun di, vy lin tc. L sp xp
hnh xon c, xp thnh 2 hng, thng, 15 - 25 2 - 3 mm, mt trn mu xanh
m hoc xn mu, mt di mu vng xanh, cung ngn; gn gia ni bt mt
trn, khng r rng mt di; thn l c 2 dy kh khng [31].
o ht c hnh cu, mu . Ht hnh trng, 6

5 mm, c 3 - 4 gc nh.

Hnh 2.14: Taxus cuspidata Siebold & Zucc [32].

A. Bi cy. B. Nn c. C. o ht trng thnh v cha trng thnh. D.Ht ging.
E. V v thn cy.
c. Phm vi phn b:
Bc Hn Quc; Trung Quc: Heilonjiang, Ct Lm, Nei Mongol, Liu Ninh,
H Bc, Thim Ty, v Sn Ty; Nga: ng v Nam lu vc sng Amur (bao
gm Sakhalin v Kurile) [26].
Nht Bn: Hokkaido, Honshu, Shikoku v Kyushu (Hartzell, 1991; Farjon,
1998) [31].

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Chng 2: TNG QUAN

Ghi ch:
Taxus cuspidata

Hnh 2.15: Phm vi phn b ca Taxus cuspidata khu vc Bc M [32].

d. Cng dng
Taxus cuspidata l mt loi cy cnh ph bin Nht Bn (Ohwi, 1965) [31]
v phng Ty k t khi du nhp vo Hoa K nm 1833.
Trc y, g cy thng c s dng ch to cc loi vt dng gia nh
nh cc loi t nh, thng cha nc, bn tm, khay, guc, a
Trong y hc, cy c s dng lm dc liu bo ch thuc ph thai v tr
bnh tiu ng. Tuy nhin, hin nay s dng gim ng k do tnh trng khai
thc qu mc (Hartzell, 1991) [31].
2.1.3.5 Taxus floridana
a. Tn thng gp
Florida yew (Hils, 1993) [24].
b. M t c im
Cy n tnh khc gc, cy bi hoc cy nh cao t 2 - 5( -10) m, ng knh
38 cm. Cnh nhiu, rm rp, phn nhnh. V cy mu nu tm, khng c vy,
mt s tui c xut hin vy mng. L mm do, c mu xanh m, 1 - 2.6( 2.9) cm

1 - 2( -2.2) mm, hnh lim, mt di mu xanh nht vi hai dy kh

khng; cung mu xanh, nh trn, gn gia khng r rt [31].

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Chng 2: TNG QUAN

o ht 1 cm

6 mm, mu ti. Ht hnh trng, 5 - 7

5 mm, thng c 2

gc nh. Ht trng thnh u ma thu.

Hnh 2.16: Taxus floridana Nutt.

A. Bi cy. B. Cnh mang nn c cha trng thnh. C. o ht.
c. Phm vi phn b
Taxus floridana c nguy c tuyt chng cao, l loi c hu ch xut hin trong
mt khu vc nh b ng sng Appalachicola qun Gadsden v Liberty, Ty
Bc Florida. Sinh trng trong iu kin m t, di tn rng, cao t 15 30 m [31].

Ghi ch:
Taxus floridana

Hnh 2.17: Phm vi phn b ca Taxus floridana khu vc Bc M [32].

24

Chng 2: TNG QUAN

d. Cng dng
Taxus floridana c cha hp cht Taxol, c hot tnh ging nh hp cht Taxol
c tm thy trong Taxus brevifolia v T. canadensis. Tuy nhin, v y l loi
qu him nn n khng c khai thc lm ngun dc liu.
2.1.3.6 Taxus globosa
a. Tn thng gp
Mexican yew (Hartzell, 1991).
b. M t c im
Cy thn g hoc cy bi, cao khong 4 - 6 m. L c mu xanh nht, ln v sc
nhn hn so vi Taxus baccata. o ht trng thnh c mu , to nht trong s
cc loi Taxus [26].

C
Hnh 2.18: Taxus globosa
A. Bi cy. B.o ht. C. Cnh mang nn c cha trng thnh.

25

Chng 2: TNG QUAN

c. Phm vi phn b
Mexico: Nuevo Len, Tamaulipas, San Luis Potos, Veracruz, Quertaro,
Hidalgo, Tlaxcala, Puebla, Oaxaca; Guatemala; Honduras; El Salvador (Hartzell,
1991) [31].

Ghi ch:
Taxus globosa

Hnh 2.19: Phm vi phn b ca Taxus globosa [34].

2.1.3.7 Taxus chinensis
a. Tn thng gp
Chinese yew.
b. M t c im [16, 28]
Cy cao khong 15 m, trong canh tc l mt loi cy bi vi tn rng, v mu
-nu i khi c mu xm. L mc tha tht trn cnh, mt s l cong ngc,
1.2 - 3 cm

2 - 3mm, mt trn l mu xanh m, mt di mu xanh-xm hoc

vng lc, nh l thay i t ngt, nhn, gn gia khng ni r mt trn ca l.

o ht hnh cu hoc gn ging hnh cu, ng knh 4 - 7mm, mu i khi
c mu xanh. Ht c hnh trng, 4 - 6

4 mm, c t 2 - 4 gc nh.

26

Chng 2: TNG QUAN

Hnh 2.20: Taxus chinensis Rehd. var mariei.

A. Bi cy. B. Cnh. C. Cnh mang o ht. D. Cnh mang nn c.
E. o ht v ht ging.
c. Phm vi phn b
Taxus chinensis mt loi c hu Trung Quc v i Loan, c tm thy
cao t 0 - 900 m. Loi ny him khi c trng Bc M, c trng nhiu
hn chu u. N c du nhp vo Anh t nm 1908.

Ghi ch:
Taxus chinensis

Hnh 2.21: Phm vi phn b ca Taxus chinensis [34].

27

Chng 2: TNG QUAN

d. Cng dng
Ch yu lm cy cnh.
2.1.3.8 Taxus wallichiana
a. Tn thng gp
Himalayan Yew, Thng Nam.
b. M t c im [32]
Cy g c kch thc trung bnh, thng xanh, cao n 20 m, ng knh thn
40 50 cm. Cnh c mu xanh nm u tin, v bin i thnh mu nu sau 3
n 4 nm. L mc xon c, gc vn nn xp thnh hai dy, hnh di di, cong
hnh ch S, di 2.5 4 cm, rng 2 3 cm, tht dn, nhn u. Cy n tnh khc
gc.
Nn c mc n c nch l. Nn ci mc n c. Ht nm trong v o
ht mu , hnh trng, c cnh di 6 7 mm.
c. Phm vi phn b
Mc ri rc trong rng rm nhit i thng xanh, ma ma m trn ni
vi.
L loi thng bn a ca vng Himalaya, t ng Afghanistan (Nepal, Bhutan,
ng Bc n (Assam, Manipur, Khasia Hills, West Bengal)) n pha Ty
Nam Trung Quc (Ty Nam Ty Tng, T Xuyn, Vn Nam), cao 2.000
3.500 m.
d. Cng dng
S dng lm cy cnh. Trong y hc, T. wallichiana l dc liu rt c gi tr,
v v l cy c cha Taxol c s dng trong iu tr ung th.

28

Chng 2: TNG QUAN

C
Hnh 2.22: Taxus wallichiana Zucc.
A. Bi cy. B. Cnh mang o ht. C. o ht v ht ging.
2.1.4 Tnh trng hin ti v bo tn loi Thng Vit Nam
Phn loi v phn b ca chi Thng (Taxus) Vit Nam khng r rng.
Danh sch Thng th gii (Farjon, 2001) (theo dng ging T. chinensis (Pilg.)
Rehder var. mairei (Leme & Lv.) W.C. Cheng & L.K. Fu nh l loi Taxus duy
nht c Vit Nam.
Thc vt ch ng Dng (Nguyn Tin Hip v Vidal, 1996) cng vi nhng
ti liu khc ghi nhn Vit Nam c hai loi: T. wallichiana Zucc. trn cao
nguyn Lm Vin min Nam Vit Nam v T. chinensis (Pilg.) Rehder ca vng
ni vi Bc v Ty Bc Vit Nam (Anon., 1996; Vu Van Dung, 1996; Nguyn
c T Lu & Thomas, 2004) (theo Nguyn Tin Hip, 2004) [1].
Cc qun th pha Bc c phn b a l gn vi ng Nam v Ty Nam Trung
Quc ni loi Taxus chinensis mc ph bin v c hnh thi gn vi cc chun
ca loi ny (T. chinensis). Cc qun th pha Nam bit lp v vic nh danh
chnh xc chng vn cha thc hin c, ph thuc vo nghin cu v phn loi
29

Chng 2: TNG QUAN

ca loi Thng Himalaya T. wallichiana, ca cc qun th Ty Nam Trung
Quc ca th T. chinensis var. mairei v ca cc qun th Indonesia v
Philippines ca loi T. sumatrana.
Hin trng phn loi khng xc nh ny cn c th hin vic cc tc gi ca
Thc vt ch Trung Quc (Fu Likuo v cng s, 1999) a th var. mairei ca
loi T. chinensis thnh mt ging thuc loi T. wallichiana.
2.1.4.1 Thng Bc (Taxus chinensis Pilg.)
a. Tn thng gp
Yew (ting Anh), hong du shan (ting Trung).
Tn ng ngha: T. wallichiana Zucc. var. chinensis (Pilg.) Florin.

Hnh 2.23 : Thng Bc (Taxus chinensis Pilg.) [1].

(V t DKH 7125 bi Nguyn Quang Hng)
1. Cnh mang nn v cc nn phn cn non. 2. L, mt di.
b. M t c im
Cy mc ng vi cc cnh tri rng (tu tng ni), t chiu cao ti 15 m
(i khi ti 20 m) vi ng knh ngang ngc ti 1 (i khi ti 1,5 m).
Cy mc ri rc trong rng nguyn sinh rm thng xanh nhit i gi ma,
ni thp trn cc dng ni vi kt tinh trng b bo mn mnh v cc sn
30

Chng 2: TNG QUAN

dc (nhit trung bnh nm 15200C, lng ma trn 1300 mm) cao 900
1500 m trn mt bin. Gp trong cc qun x Thit sam gi (Pseudotsuga
sinensis), Thng P C (Pinus kwangtungensis), P mu (Fokienia hodginsii) hoc
Bch vng (Xanthocyparis vietnamensis), xen vi Thng tre l ngn (Podocarpus
pilgeri), Kim giao ni (Nageia fleuryi) v Thit sam ni (Tsuga chinensis).
Thnh thong c ti sinh t nhin [1].
c. Phm vi phn b
Thng bc gp cc tnh Sn La, H Giang, Tuyn Quang, Cao Bng, Ho
Bnh v Thanh Ho [1]. Trn th gii loi cn gp Nam Trung Quc.

Hnh 2.24: Phm vi phn b ca Thng Bc

(Taxus chinensis Pilg.) Vit Nam [1].
d. Cng dng
G Thng Bc c mu vi cu trc mn, chu ngm v c th s dng
lm cc gung tt nc. Ht c tnh cht cha bnh v cy c dng lm cy th.
Vit Nam nghin cu v dc tnh ca loi ny cn hn ch. Nhng hot
cht quan trng sn xut Taxol nh 7xylosyl10deacetylbaccatine III v 10
deacetylbaccatine III c phn lp v m t t v v l cy Thng Bc
(Taxus chinensis) Ho Bnh (Mai Vn Tr v cng s, 1995) (theo Nguyn Tin
31

Chng 2: TNG QUAN

Hip, 2004) [1]. Vic thu hi ch c th bn vng nu cy c trng v cc qun
th t nhin u qu nh.
e. Hin trng bo tn [1]
Hin trng quc t

CHA NH GI [T LIN QUAN]

Hin trng quc gia hin ti

HIM

Hin trng quc gia qua xut nh gi

SP TUYT CHNG

Do loi c phn b rng Nam Trung Quc nn mc quc t khng c coi

l b e do tuyt chng. Vit Nam kch thc cc qun th u nh hn rt
nhiu, v nhiu qun th b cht h. mc quc gia loi ny c coi l
Him. Theo cc tiu ch mi ca IUCN (2001) mc nh gi ca loi ny cn c
chuyn thnh Sp b tuyt chng.
Cc mi e da
Sinh hc sinh sn phc tp v kh nng ti sinh km ca Thng Bc i lin
vi nhng e do b cht h, rng b chia ct, ni sng b suy gim v chy rng.
S hin din trong cc khu bo tn
Cc qun th nh Thng Bc Ho Bnh c thy khu bo tn t nhin
Hang Kia P C. Qun th ny c th l mt phn ca qun th Sn La, ni cha
c cc khu bo tn (Thomas v Nguyen Duc To Luu, 2004c) [1].
H Giang, Thng Bc c bit c khu bo tn t nhin Bt i Sn,
Tuyn Quang c trong khu bo tn t nhin Na Hang. Cy cn c tm thy
trong khu bo tn xut P Lung.
Cc bin php bo tn thc hin
Cha c nhng th nghim bt k qui m no nhm thun dng Thng
Bc min Bc Vit Nam. Mt s nghin cu phng php nhn ging c Vin
Khoa Hc Lm Nghip Vit Nam (Nguyen Hoang Nghia, 2000) [1] thc hin.
Thng Bc v cc loi Thng chu khc c a vo Ph lc II ca
Cng c CITES.
Hot ng bo tn xut

32

Chng 2: TNG QUAN

Thng Bc cn c a vo Nhm IIA ca Danh mc cc loi ng vt v
thc vt qu him nhm to khung php l cho vic bo v cc khu vc c cc
qun th ch yu v km theo vic o to cc cn b kim lm a phng.
Nghin cu phn loi tip theo l cn thit nhm lm r v tr phn loi ca loi
. Vic bo tn chuyn v cng cn c thit lp.
2.1.4.2 Thng Nam (Taxus wallichiana Zucc.)
a. Tn thng gp
Thng l di (ting Vit), Himalayan Yew (ting Anh), xu mi hong dou
shan (ting Trung).
Tn ng ngha: T. yunnanensis W.C. Cheng & L.K. Fu.

Hnh 2.25: Thng Nam (Taxus wallichiana Zucc.) [1].

(V t DKH 7125 bi Nguyn Quang Hng)
1. Cnh mang nn v cc ht chn. 2. L, mt di.
b. M t c im
Cy g mc ng, n tnh khc gc vi cc cnh tri rng tu theo a im,
t chiu cao ti 20 (i khi ti 30) m vi ng knh ngang ngc ti 1 (i khi
1,5) m. Cy mc thnh cc qun th nh cc b trong rng nguyn sinh rm
thng xanh hn giao hoc cy l rng chim u th nhit i gi ma ni thp
(nhit trung bnh nm 15 21 C, lng ma trn 1500 mm) trn t phong ho
t granite hoc cc m silicat khc cao 900 1600 m.
33

Chng 2: TNG QUAN

Cc loi Thng mc cng gm nh tng (Cephalotaxus mannii), Kim giao ni
t (Nageia wallichiana), Thng tre l di (Podocarpus neriifolius), Thng nng
(Dacrycarpus imbricatus) v Du sam ni t (Keteleeria evelyniana). Ti sinh t
nhin khng thng xuyn, cy m v cy con chu bng tt.
c. Phm vi phn b
Vit Nam Thng Nam hin ch c bit chc chn c Lm ng (c
Trng, Xun Th, n Dng, Lc Dng) [1]. C cc mu vt kh c t nhng
nm 1930 ghi nhn loi ny c Khnh Ho (Nguyen Tien Hiep v Vidal, 1996)
[1] nhng khng c cc thng tin mi t cc im ny. Nhng thng tin c Ninh
Thun v c Lc cn c kim tra thm. Trn th gii loi ny c thy Ty
Nam Trung Quc v dy Himalaya.

Hnh 2.26: Phm vi phn b ca Thng Nam (T. wallichiana Zucc.)

Vit Nam [1].
d. Cng dng
Nghin cu Vit Nam v dc tnh ca Thng Nam cn hn ch. Mt
phn tch cc mu cy thu t Lm ng cho thy n c hm lng 10
deacetylbaccatine III cao hn loi Taxus baccata (L Th Xun v cng s, 1996)
(theo Nguyn Tin Hip, 2004) [1].

34

Chng 2: TNG QUAN

e. Hin trng bo tn [1]
Hin trng quc t

THIU THNG TIN [T LIN QUAN]

Hin trng quc gia hin ti

HIM

Hin trng quc gia qua xut nh gi

ANG B TUYT CHNG

Do c phn b rng trn th gii nn Thng Nam hin nay cha c coi l
b e do tuyt chng mc d nhiu qun th b khai thc qu mc.
Vit Nam loi ny c nh gi l loi Him (Anon., 1996), Thiu thng
tin hoc ang b tuyt chng trm trng (C2a) (Nguyen Hoang Nghia, 2000) [1].
nh gi sau cng da trn nhng iu tra hin trng ti Lm ng t nm
1998 m c tnh tng kch thc qun th t hn 250 cy trng thnh vi mi
tiu qun th khng c qu 50 cy. T ti nay, nhng iu tra hin trng tip
tc c tin hnh Lm ng, v mt qun th c t nht 250 cy c pht
hin. Tng s cy trng thnh Vit Nam hin nay c tnh nhiu hn 250 c
th, nhng chc chn t hn 2500 cy, mc ri rc thnh cc qun th b chia ct
mnh [1].
Nh vy, hin trng ca loi ny Vit Nam cn c nh gi mc ang b
tuyt chng (C1) theo cc tiu ch ca IUCN (2001). V tt c cc qun th u i
mt vi nhng e do ln, c bit l chy rng t cc khu vc ln cn nn hin
trng bo tn ca loi ny c th phi c nng ln trong tng lai. Cc qun th
Lm ng l im phn b cc nam ca chi Thng lc a ng Nam .
Do cc qun th ny phn b bit lp nn c th l mt xut x ring bit (Thomas,
Gardner v Nguyen Duc To Luu, 2002) [1].
Cc mi e da
Nhn chung Thng Nam b e do tuyt chng do mt cc ni sng, c th
l do chy rng cc khu rng ln cn. Vic bun bn cc b phn hoc cc cht
chit ca loi ny lm ngun ch Taxol l e do i vi cc qun th t nhin
nhiu ni phn b trn th gii (Thomas v Nguyen Duc To Luu, 2004c) [1].
S hin din trong cc khu bo tn
Nam Vit Nam tt c cc qun th Thng u nm trong cc khu vc
rng c dng nhng khng phi l cc khu vi mc ch bo tn a dng sinh

35

Chng 2: TNG QUAN

hc. Hai qun th chnh Ni Voi (c Trng) v H Tin (n Dng) l rng
phng h u ngun c qun l bo v.
Qun th th ba Xun Th ( Lt) nm trn t X nghip ging lm nghip
qun l v l rng th nghim (rng ging). Ni B p l khu Bo Tn Thin
Nhin nhng y mi ch quan st thy c 1 c th, tuy vn c nhiu kh nng
tm thy nhng cy khc (Thomas v Nguyen Duc To Luu, 2004c) [1].
Cc bin php bo tn thc hin
Loi ny c bo v bi lut php t khi c xp vo Nhm IA ca Danh
lc cc loi ng vt v thc vt qu him. Vin KHLN Vit Nam (Nguyen Hoang
Nghia, 2000) v Cng ty ging LNT (Dick v cng s., 2004) tin hnh nhn
ging sinh dng v trng bo tn chuyn v . Taxus wallichiana cng vi cc loi
Thng chu khc c a vo Ph lc II ca Cng c CITES.
Hot ng bo tn xut
Thng Nam Ni Voi, c Trng, Lm ng cn c ngh l mt khu
bo tn (v d nh khu bo tn ni sng cho loi) i vi Thng, c bit l Thng
Nam (Taxus wallichiana). Do loi ny c php lut bo v nn cn nng cao
nhn thc cho cc c quan lin quan v nhn dn nhng a im c cy cn li
nhm mc ch bo tn lu di. Qun th ny cng l a im tt cho nghin cu
v sinh thi v ht ging cho Thng Nam. Cn tip tc cc nghin cu v phn
loi nhm lm r v tr phn loi ca loi.
2.2 Taxol
2.2.1 Lch s nghin cu
Hp cht diterpenoid Taxol l mt trong nhng hp cht t nhin c ngun gc
t thc vt c kh nng chng ung th hiu qu nht hin nay. Taxol cng l loi
thuc iu tr ung th c gi tr thng mi hng u th gii, c tnh doanh thu
hng nm trn 3 t USD trn ton th gii [11].
Taxol cho hiu qu trong iu tr ung th bung trng v ung th v v cho
thy trin vng trong iu tr mt s loi bnh ung th khc nh no, c, phi v
bng quang (Holmes v cng s, 1995; Suffness, 1995) [11].
Taxol ban u c chit sut t v cy Taxus brevifolia Nutt. Cu trc Taxol
c xc nh bi Wall v Wani v cng s vo nm 1971 (Wani v cng s,
1971) [29] v c chp thun thnh sn phm thng mi ln u tin bi Cc
36

Chng 2: TNG QUAN

qun l Thc phm v Dc phm Hoa k trong iu tr ung th bung trng vo
nm 1992 v iu tr di cn ung th v vo nm 1994 (Suffness v Wall, 1995)
(theo Croteau v cng s, 2006) [14].
Thi gian pht trin lu di ca loi thuc ny do nhiu nguyn nhn: ngun
gc cung cp hp cht ny t Taxus brevifolia (Pacific yew) cn hn ch (vic thu
hoch lm tn ph ngun cung cp t nhin, qu trnh tch chit phc tp, sn
phm thu c khng cao, nhng ngc li hiu qu iu tr cao), ngoi ra cn do
tnh cht k nc ca cc nhm diterpenoid t nhin v cc tc dng ph trong th
nghim iu tr lm sng (Arbuck v Blaylock, 1995) [11].
C ch tc ng ca Taxol l kh mi l v c o. Tri ngc vi Vinca
alkaloid, cochicine v mt s cht chng ung th khc vi c ch tc ng l ngn
cn s trng hp ca tubulin vo trong vi ng (microtubule), Taxol y nhanh qu
trnh lp rp ca microtubule v ngn cn qu trnh depolymerization (kh trng
hp) (Horwitz, 1992; Jordan v Wilson, 1995; Arbuck v Blaylock, 1995) [22].
Vic thu hoch Taxus sn xut Taxol c nguy c dn ti ph hoi mi
trng t nhin v sn lng Taxol thu c qu thp khng cung cp cho nhu
cu iu tr ngy mt tng. Do , nhiu phng php thay th ngun cung cp
Taxol c nghin cu.
Mc d, mt vi nhm (Cragg v cng s, 1993; Holton v cng s, 1994;
Nicolau v cng s, 1994, 1995; Masters v cng s, 1995) [11] t c kt
qu tng hp Taxol bng phng php ha hc nhng chi ph cao ca phng
php ngn cn kh nng thng mi ha ca n.
Taxol c cng c sn xut bng phng php bn tng hp t tin cht
mi (v d: baccatin III) c th d dng thu nhn t cc loi cy l kim khc nh
mt ngun cung cp nguyn liu c trin vng.
Mt phng php tip cn khc l s dng phng php nui cy dch treo t
bo ca T. brevifolia. Mt vi nhm nghin cu t c thnh cng trong sn
xut Taxol hoc bacatin t dch treo t bo Taxus (Yukimine v cng s, 1996;
Hezari v cng s, 1997) [11].
Taxol cng c tch chit t v, l, cnh v tri ca cy ph (hazelnut) nhng
sn lng thu c ch bng 10% so vi t cy Thng . Gn y, mt loi nm
Taxomyces vreanae c tm thy c kh nng sn xut Taxol v kt qu cng
37

Chng 2: TNG QUAN

tng t nh Taxol thu c t Thng (Stierle v cng s, 1993; Strobel v
cng s, 1996).
Bng 2.1: Tm tt lch s nghin cu v hp cht Taxol (Wall v Wani, 1998) [25].
Thi
gian

S kin
Hot ng chng ung th trong cc hp cht chit sut t v cy T.

1963

brevifolia c khm ph trong mt cuc kim tra quy m ln trn

35000 loi thc vt.

1971

Taxol c xc nh l hp cht c kh nng chng ung th.

1979

C ch duy nht ca Taxol c xc inh l hot ng lin kt to s n

nh cu trc ca tubulin.

1983

NCI tin hnh th nghim lm sng v s an ton v hiu qu ca Taxol

chng li nhiu loi ung th.

1989

Taxol p ng nhu cu iu tr mt phn hoc hon ton 30% bnh nhn

c iu tr trc vi ung th bung trng.

1991

Cng ty BristolMyers Squibb c chn l i tc ca NCI trong vic

pht trin tim nng ca TAXOL v k kt CRADA vi NCI

1992

FDA chp thun vic s dng Taxol trong iu tr ung bung trng. Tin
hnh th nghim lm sng chng minh tc dng ca thuc trong iu tr
ung th v di cn.

1993

Cng ty BristolMyers Squibb kt thc qu trnh thu hoch v cy T.

brevifolia v bt u pht trin cc ngun cung cp nguyn liu sn xut
Taxol khc.
FDA chp thun vic s dng Taxol trong iu tr ung th v di cn, nu

1994

bnh nhn ti pht trong vng 6 thng k t khi dng phng php ha tr
liu hoc khng p ng vi phng php ha tr liu kt hp.

1995

Dng bn tng hp ca Taxol c FDA chp thun a vo th trng.

1997

FDA chp thun dng bn tng hp ca TAXOL trong iu tr AIDS giai

on 2, lin quan ti Kaposis sarcoma. Taxol sn xut bng phng php
nui cy t bo c ph chun ti Hn Quc v c xut khu nh l sn
phm Genexol ca Samyang Genex.

1998

FDA chp thun cho dng bn tng hp ca TAXOL kt hp vi cisplatin

trong giai on u iu tr ung th phi cho nhng bnh nhn khng th
iu tr bng phu thut hoc x tr.
38

Chng 2: TNG QUAN

2.2.2 Cu to Taxol
Trong cc hp cht th cp c dng trong iu tr ung th, Taxol l mt
diterpene phc tp thu c t Taxus spp., c cho l quan trng nht.
Taxol i din cho khong 400 nhm taxoid c xc nh (v d, taxane
diterpenoid) ca Taxus spp., tt c u da trn mt b khung taxane duy nht
(pentamethyl [9.3.1.0] tricyclopentadecane) hoc do s sp xp li sn phm ca
khung tricyclic (v d, abeo-taxoids c to thnh do s sp xp v tr A/B hoc
B/C trn vng). Cc nhn taxane mang 3 nhn lp th (stereocenter) v bn thn
Taxol mang 11 trung tm nh vy, chnh s lng ln nhn lp th nh vy dn
ti nhng kh khn nht nh trong tng hp Taxol [27].
Tn ha hc ca Taxol l 5, 20-epoxy-1,2,4,7,13-hexahydroxytax-11-en9one-4,

10-diacetate-2-benzoate

13

ester

vi

(2R,3S)-N-benzoyl-3-

phenylisoserine.
Cng thc phn t C47H51NO14 v khi lng phn t 853.9 Da.
Ti nhn ca Taxol l h thng vng A, B v C, vi mt s nhm chc bao
gm hai nhm OH, mt nhm benzoyl, hai nhm acetyl v mt vng oxetane. Lin
kt vi C13 ca nhn l mt chui bn hoc C13 (2R,3S)-N-benzoyl-3phenylisoserine vi mt nhm OH v mt nhm benzoyl.

Hnh 2.27: Cu trc ca Taxol [20].

39

Chng 2: TNG QUAN

2.2.3 Con ng sinh tng hp Taxol
S gia tng nhu cu s dng kt hp Taxol trong iu tr ung th, v ngun
cung cp hn ch trong t nhin i hi phi c mt s hiu bit cn k v c ch
sinh tng hp ca n, c bit l c ch phn ng ca cc enzyme.
2.2.3.1 Cc nghin cu trc v qu trnh sinh tng hp Taxol [11]
Cu trc ca Taxol l kh phc tp c mang mt b khung C diterpene khc
thng, tm nhm oxy chc nng v phi hp vi mt chui acyl, vi 11 trung
tm phn ng (Wani v cng s, 1971) [29]. Theo xut ban u ca Harrison v
cng s (1966) [11], qu trnh sinh tng hp Taxol bt u vi s hnh thnh b
khung olefinic taxane, tip sau l mt lot cc phn ng oxy ha, cui cng l
phn ng acyl ha (Zamir v cng s, 1992; Eisenreich v cng s, 1996) [11].
Da trn cc nghin cu v s trao i cht ca cc nhm terpenoid khc
(Gershenzon v Croteau, 1993) [11], con ng sinh tng hp Taxol cng c
bt u t tin cht chung ca diterpenoid l GGPP lin quan ti hn mi phn
ng enzyme. Gii m qu trnh sinh tng hp Taxol t ra mt thch thc ng k
v cu trc phc tp ca n, nng sut thu nhn thp v qu trnh th nghim kh
khn i vi cc loi Taxus.
Cc nghin cu trc y, c thc hin in vivo vi cc tin cht c nh
du ng v phng x cho thy, acetate, mevalonate, v phenylalanine xy dng
nn cu trc ca Taxol (Zamir v cng s, 1992) [11]. Tuy nhin, gn y
Eisenreich v cng s (1996) ch ra rng h thng vng taxan c hnh thnh
t con ng c lp vi con ng mevalonate c tm thy bi Rohmer trong
sinh tng hp isoprenoid (Schwender v cng s, 1996) [11].
Nm 1996, Eisenreich v cng s trong bo co th nghim vi tin cht c
nh du ng v phng x 13C trong dch treo t bo ca Taxus chinensis. Phng
php ny to ra 2.6 % diterpene taxuyunnanine C trong sn phm (da trn trng
lng kh). Cc mu ng v phng x chng minh taxuyunnanine C c tng
hp thng qua con ng ngoi mevanlonate.
2.2.3.2 Con ng sinh tng hp Taxol
Qu trnh sinh tng hp Taxol trong chi Taxus cha c khm ph y .
Terpenoid l mt nhm hp cht th cp c cu to rt a dng vi tham gia cu
trc v chc nng trong thc vt. Mt s terpenoid ng vai tr thit yu i vi 1
40

Chng 2: TNG QUAN

s loi thc vt, v d nh phytohormone (hormone thc vt), cc sc t quang hc
hoc tham gia thnh phn cu trc mng t bo, trong khi cc nhm khc tham
gia trong cc phn ng phng v v thch ng ca thc vt vi mi trng sng.
Tuy nhin, tt c chng u c hnh thnh t cng mt tin cht, isopenthenyl
diphosphate (IPP) v dimethylallyl diphosphate (DMAPP) [18].
Trong nhiu loi thc vt, c s chia i trong con ng sinh tng hp cc
tin cht ca terpenoid.
Trong t bo cht, con ng mevalonate sn xut IPP t acetyl coenzyme A
sinh tng hp, sterol, triterpene v mt s sesquiterpene. Cc lp th l ni din
ra con ng thay th ngoi mevalonate sn xut IPP v DMAPP t pyruvate v
glyceraldehyde-3-phosphate to thnh cc isoprenoid: monoterpene, diterpene v
tetraterpene. Theo Eisenreich v cng s (1996), cc nghin cu v sinh tng hp
Taxol kt lun h thng vng taxoid khng c tng hp qua con ng
mevalonate., iu ny ph hp vi cc diterpene t nhin. Tuy nhin, mt s
nghin cu trc thc hin vo nm 1991, cho rng h thng vng taxoid c
tng hp t con ng mevalonate. ng thi, kt qu khi nui cy t bo T.
media (T. cuspidata x T. baccata) c b sung mevalonate cho thy c s tham gia
ca c hai con ng.
Cc kin hin ti u chp nhn rng, mt s t bo Taxus s dng con
ng mevalonate l ngun cung cp IPP sinh tng hp Taxol, v tin cht
chnh tng hp terpenoid l GGPP t con ng ngoi mevalonate.
Bc phn ng u tin trong con ng sinh tng hp Taxol l s to vng
ca geranylgeranyl diphosphate (GGPP) thnh taxa-(4,5),(11,12)-diene, mt phn
ng xc tc bi taxadiene synthase (TS), mt protein monomer 79kDa. Enzyme
ny c tinh ch v m t bi Hezari v cng s, v gen m ha cho TS c
to dng v biu hin chc nng trong E.coli bi Wildung v Croteau.
Sau , oxy v cc nhm acyl c b sung thm vo nhn taxane bi phn
ng oxy ha nhiu v tr trung gian bi cytochrome P450 mono-oxygenase.
Phn ng hydroxyl ha ti v tr C5 ca vng taxane c xc tc bi enzyme
cytochrome P450 taxadiene-5-hydroxylase (T5H), kt qu dn ti s hnh thnh
ca taxa-4(20),11(12)-dien-5-ol, l bc th 2 trong sinh tng hp Taxol. T5H
l mt protein c khi lng 56 kDa vi trnh t N cui cng, enzyme ny c

41

Chng 2: TNG QUAN

gn trn mng ca mng li ni cht. Enzyme ny ngoi hot ng xc tc cho
phn ng hydroxyl ha, n cng xc tc chuyn v ni i t 4(5) thnh 4(20).
Mc d 2 bc chuyn ha, to vng v hydroxyl ha din ra chm, nhng n
khng hn ch t l trong sinh tng hp Taxol.
Bc tip theo trong con ng sinh tng hp c xc tc bi taxadiene-5ol-O-acetyl transferase (TDAT), acyl ha taxa-4(20),11(12)-dien-5-ol thnh taxa4(20),11(12)-dien-5-yl-acetate. Enzyme ny c khi lng 50kDa khng mang
trnh t N cui cng. Cc sn phm ca phn ng ny sau c hydroxyl ha
bi taxoid 10-hydroxylase (T10H) ti C10. T10H l mt P450-dependent
monooxygenase c to dng v c biu hin c im trong nm men.
Mt Cyt P450-dependent hydroxylase dn ti s hnh thnh ca taxa4(20),11(12)-dien-5-13-diol c tm thy. Thc t l enzyme ny s dng
c cht tng t nh (TDAT), taxa-4(20),11(12)-dien-5-ol, cho thy qu trnh
sinh tng hp Taxol khng phi ch c duy nht mt con ng m c nhiu im
phn nhnh dn ti s hnh thnh cc nhm taxoid khc c lin quan. Trng hp
thay th ny c quan st, c bit n thng xy ra trong qu trnh nui cy
t bo vi s c mt ca methyl jasmonate.
Taxoid 14-hydroxylase (T14H) xc tc s hnh thnh ca taxa-4(20),11(12)dien-5-acetoxy -10-14-diol. Enzyme ny khng s dng c cht c
hydroxyl ha ti v tr C13, m ch nhng c cht c hydroxyl ha ti v tr C10,
cho thy T14H khng th tham gia vo vic sn xut Taxol, m cng khng xut
hin phn ng hydroxyl ha ti v tr C14.
Bc cui cng trong con ng sinh tng hp Taxol, sau khi hnh thnh taxa4(20),11(12)-dien-5-yl-acetate, bao gm 1 s v tr c hydroxyl ha ti C1,
C2, C4 v C7, oxy ha ti v tr C9, v epoxide ha ti v tr ni i C4, C5. Phn
ng hydroxyl ha c xc tc bi cc enzyme Cyt P450 nhng n khng chnh
xc trong mt s trng hp.

42

Chng 2: TNG QUAN

Hnh 2.28 : Con ng sinh tng hp Taxol bt u vi geranylgeranyl diphosphate

v kt thc vi tin cht gi nh c polyhydroxyl ha [18].
Mc d, c nhiu c ch khc nhau cho s hnh thnh vng oxetane c
xut, tuy nhin hin ti c ch c chp nhn l: qu trnh ny c lin quan n
qu trnh epoxide ha ti v tr ni i 4(20) vi s chuyn v ca nhm -acetoxy
t v tr C5 n C4 cng vi vic m rng nhm oxirane thnh nhm oxetane.

43

Chng 2: TNG QUAN

C th bc ny trc bc hydroxyl ha ti v tr C1 trong sinh tng hp
Taxol, v trong trng hp ny gi nh polyhydroxyl ha l cht trung gian s l
taxadien-hexaol hn l heptaol hydroxyl ha ti C1. Enyme xc tc phn ng
epoxide ha ti ni i C4-C20, v c ch m rng vng oxirane thnh oxetane
cng cha c bit n y .

Hnh 2.29: Qu trnh hnh thnh vng oxetane trong sinh tng hp Taxol [18].
Sau khi hnh thnh cc tin cht gi nh c polyhydroxyl ha
(hypothetical polyhydroxylated precursor), n c xc tc phn ng tip tc bi
2-O-benzoyl transferase (DBT), protein c khi lng 50 kDa, hp cht thu c
tip theo l 10-deacetylbaccatin III. Mt phn ng chuyn nhm acetyl c xc
nh trong con ng sinh tng hp Taxol lin quan ti phn ng hydroxyl ha ti
v tr C10 ca 10-DAB (10-deacetylbaccatin III) vi s xc tc ca enzyme 10deacetyl-baccatin III-10-O-acetyl transferase (DBAT). Dn ti s hnh thnh mt
diterpene trung gian, baccatin III, s dng c cht l 10-DAB v acetyl CoA.
Mt bc cn thit trong sinh tng hp Taxol l phn ng ester ha ca nhm
hydroxyl C13 ca baccatin III vi phenylalanoyl-CoA ca chui bn. Cc chui
bn hnh thnh t cc acid amin- phenyalanine bi tc ng ca phenylalanine
aminomutase (PAM). Mt ester CoA ligase cha c bit n c th kch hot
hp cht ny v n c th lin kt vi cc baccatin III. Enzyme xc tc cho phn
ng lin hp ca chui bn phenylalanoyl-CoA v baccatin III l C-13phenylpropanoyl-CoA transferase (BAPT), sn phm l cc hp cht 3-Ndebenzoyl-2-deoxytaxol. Hp cht ny c chuyn ha bi enzyme Cyt P450dependent hydroxylase c ch cha c bit r, xc tc phn ng hydroxyl v
tr

C2

enzyme

3-N-debenzoyl-2-deoxytaxol

N-benzoyl

transferase

44

Chng 2: TNG QUAN

(DBTNBT) xc tc phn ng lin hp vi benzoyl-CoA thnh 3-N-debenzoyl-2deoxytaxol, Taxol l hp cht cui cng.

Hnh 2.30: Con ng sinh tng hp Taxol t tin cht gi nh c

polyhydroxyl ha [18].
2.2.4 C ch tc ng ca Taxol
Hot ng chng ung th ca Taxol ch yu do chui mch bn, vng A, nhm
benzoyl C2 v vng oxetane. Hot ng ny c duy tr bi nhm amide-acyl
vi tr C3 trong chui gn v tr C13 v c tng cng bi nhm hydroxyl v
tr C2 [27].
45

Chng 2: TNG QUAN

S tng tc ca cc thnh phn trn vi -tubulin ca vi ng, thc y qu
trnh polymer ha sn xut cht c v n nh vi ng.
Taxol c ch s pht trin t bo bng cch lin kt vi b mt ca vi ng, c
th l lin kt vi tiu n v ca tubulin l cc heterodimer, do thc y qu
trnh trng hp ca n, ngay c trong trng hp khng c GTP.

Taxol
Taxol

Qu trnh polymer ha ca tubulin

n nh cu trc vi ng
(c ch phn ng depolymerization)

T bo ung th khng thc hin c cc

hot ng thng thng

Chu k phn chia t bo ngng giai on G2/M

T bo ung th cht hng lot

Hnh 2.31: C ch tc ng ca Taxol i vi t bo gy ung th [25].

46

Chng 2: TNG QUAN

Vi ng bao gm cc polymer ca tubulin heterodime trng thi cn bng
ng. Tubulin heterodime gm c cc protein tiu n v v . Vi ng c hnh
thnh t s nh trng hp 2 loi tubulin l v . Mi loi tubulin c khng 450
loi acid amin, s tng ng v acid amin gia chng l khong 40%, v chng
c cu trc tng t nhau. Mi monomer ny c cu trc xon v gp np . N
cng bao gm mt khu vc lin kt vi nucleotide, lin kt vi cc protein khc v
3 v tr lin kt vi Taxol [18, 25].
C mt trng thi cn bng ng lin tc gia tubulin nh trng hp v vi ng,
do c s cn bng gia qu trnh trng hp v kh trng hp
(depolymerization). Tc ng ca Taxol ti a khi t l gia Taxol/tubulin l 1/1 v
qu trnh polymer ha ca tubulin c thc y khi nng ca Taxol l 0.05
mM. Lin kt ca Taxol vi cc vi ng s khng thay i, vi hng s lin kt vo
khong 0.9 mol/L [18].
Mc d, chc nng ch yu l hnh thnh thoi v sc trong qu trnh phn chia
t bo, vi ng cng tham gia nhiu chc nng quan trng k trung gian bao gm
duy tr hnh dng, vn ng, dn truyn tn hiu v vn chuyn ni bo.
Khng ging nh cc loi thuc ngn cn s hnh thnh vi ng, nh vinca
alkaloid, gy ra s tch ri ca vi ng, Taxol li thc y qu trnh trng hp ca
tubulin. nng nh hn 10-9, Taxol c ch s tch ri ca vi ng, trong khi
n lm tng s lng v khi lng ca chng, y l nng trong iu tr lm
sng.
Cc vi ng hnh thnh khi c mt Taxol cc k n nh v gy ra ri lon chc
nng, do lm cht t bo bng cch ph v trng thi cn bng ng bnh
thng ca qu trnh phn chia t bo cng nh cc qu trnh chun b k trung
gian. Taxol cng gy ra nhng biu hin trn gen i vi nhn t gy ra khi u
hoi t, nhng nghin cu v cu trc v hot ng cho thy hot ng ny khng
lin quan ti tc ng ca Taxol i vi s lp rp vi ng, qu trnh nui cy m
cho thy mt phn vai tr ca cytokines trn hot ng khng ung th ca Taxol.
V tr lin kt ca Taxol l khc bit so vi v tr lin kt ca guanosine
triphosphate, colchicine, vinblastine, v podofilox (podophyllotoxin). Taxol lin
kt vi N-teminal 31 amino acid ca -tubulin, nhiu hn l lin kt vi tubulin
nh hp gia v .

47

Chng 2: TNG QUAN

Hnh 2.32: C ch phn ng ca Taxol trong h thng vi ng ca t bo [18].

Hai c ch khng thuc i vi Taxol:
C ch th nht, mt s khi u c cha c v -tubulin lm gim
kh nng tc ng ca Taxol vo qu trnh polymer ha cng nh lm chm tc
lp rp ca vi ng.
C ch th 2 lin quan ti s khuych i ca phn t
phosphoglycoprotein trn mng, n c chc nng nh mt my bm thuc ra
ngoi.
Trong cc t bo nguyn vn, Taxol to thnh tng b vi ng, v tc ng ti
s hnh thnh mt s lng ln thoi v sc. Taxol cng tng cng hot ng khi
c s tc ng ca bc x ion in vitro, bng cch ngn chn trong giai on G2 ca
k trung gian v trong qu trnh phn bo l nhng giai on nhy vi bc x.
Tnh kh thi ca vic s dng Taxol kt hp vi bc x iu tr cho bnh
nhn ung th phi, u v c, ung th thc qun c th p ng vi c hai phng
php iu tr hin ang c nh gi.

48

Chng 2: TNG QUAN

Trong khi s lng bnh ung th c iu tr bng Taxol ngy cng c m
rng, cho n nay n c ch yu dng trong iu tr ung th bung trng, ung
th v di cn v mt s lng nh trong iu tr ung th phi cng nh iu tr
AIDS giai on 2-lin quan ti ung th m lin kt (Kaposis sarcoma).
Taxol hin ang c nghin cu iu tr cc bnh khc do n c kh nng
n nh vi ng cng nh ngn cn s pht trin ca t bo v s hnh thnh mch,
v d nh bnh psoriasis (bnh vy nn).
Taxol cng ang c nghin cu iu tr taupathies (tc ng ti cc
protein tau-protein n nh vi ng, chng c nhiu t bo thn kinh trong h thn
kinh trung ng), chng hn nh bnh Alzheimer, hi chng Parkinson lin qun
ti nhim sc th s 17, v mt s bnh khc.
2.2.5 Mt s phng php tng hp hp cht Taxol
2.2.5.1 Tng hp bng phng php ha hc
Cu trc ca Taxol, ngoi b khung chnh: vng A, B v C, n cn bao gm
vng oxetane (vng D), chui bn N-benzoyphenylisoserine ni vi C13 ca vng
A v nhm benzoate v tr C2 ca vng B.
Nm 1994, hai nhm nghin cu (Holton v cng s, 1994; Nicolaou v cng
s, 1994) [11] bo co v hp cht Taxol tng hp bng phng php ha hc
vi mt t l thp, tng ng l 2.7% v 0.07% [21].
Theo nhm ca Nicolaou v cng s, phng php tng hp c s dng l
xy dng cc vng A v C ring bit sau ni 2 phn t li vi nhau bng cch
s dng phn ng Shapiro v lin kt McMurry hnh thnh vng B. Tip theo
sau l mt lot cc phn ng gn vng oxetane (vng D) v cc nhm chc
bn ngoi ca vng B v C. cui giai on, l qu trnh oxy ha v ester ha v
tr C13 gn chui bn v hon thnh cu trc Taxol.

49

Chng 2: TNG QUAN

Hnh 2.33: Phng php phn tch retrosynthetic ca Nicolaou [11].

Holton v cng s, c mt cch tip cn khc, phng php a ra l s dng
(-)-borneol l vt liu khi u, sau c bin i thnh ketone khng bo ha
vi khong trn 13 bc tng hp.

50

Chng 2: TNG QUAN

Hai nm sau, Danishefsky v cng s (1996) xut con ng tng hp th
ba vi cc bc tng hp t hn so vi 2 phng php ca Nicolaou v Holton.
Phng php c khi u vi Wielv-Miescher ketone, tip c bin i
thnh hp cht enol triflate c mang olefin trn vng C cho php pht trin thnh
Taxol thng qua phn ng Heck ni phn t.
Ngoi ra, nm 1998 Morihira v cng s bo co tng hp thnh cng hp
cht Taxol. Mc d, tng hp thnh cng hp cht Taxol bng phng php ha
hc l mt thnh tu khoa hc ln, nhng n khng th c thng mi ha
trong tng lai gn v cc phng php ny cn t nht 20 bc tng hp ha hc
v nng sut mt s bc l tng i thp. iu ny lm cho phng php ny
rt phc tp v tn km nhiu chi ph.
2.2.5.2 Nui cy t bo Taxus
Nhu cu i vi vic cung cp Taxol trong iu tr ung th ngy mt tng, y
l kt qu ca vic s dng m rng Taxol trong cc liu php can thip sm cng
nh kt hp ci cc tc nhn ha tr liu khc.
Do nhng vn bao gm: khai thc t cc ngun t nhin v nhng kh khn
ca ca phng php bn tng hp, c nhng nghin cu thu nhn Taxol
da trn nui cy t bo. Cng ngh sinh hc ng dng trong sn xut Taxol c
nghin cu t nhng nm u thp nin 90. Ban u cng vic c thc hin bi
nhiu nhm nghin cu khc nhau cho thy m so v dch treo t bo t thn cy
non ca Taxus sp. c th sn xut Taxol t nht l mc tng t nh cy
trng.
tng nng sut ca Taxol v cc taxoid c lin quan trong nui cy m v t
bo, c th s dng nhiu phng php khc nhau bao gm: ti u ha cc iu
kin nui cy, la chn dng t bo c nng sut cao, s dng cht cm ng
(elicitor), v b sung tin cht, cc phng php ny c kim tra bi nhiu
nh nghin cu. y l nhng bo co nh gi cng ngh sinh hc trong sn
xut Taxol v cc kin thc hin ti v s trao i cht ca Taxol trong nui cy t
bo.
Bng 2.2 tm tt cc nghin cu quan trng gn y trn kh nng nng cao
sn xut Taxol bng k thut nui cy t bo v phng php chnh c s dng
t c mc tiu ny.
51

Chng 2: TNG QUAN

Bng 2.2: Cc nghin cu gn y v kh nng thu nhn Taxol v cc hp cht
c lin quan bng phng php nui cy t bo [18].
Tm tt nghin cu

Tc gi

Mt dng bioreactor mi c pht trin

thnh cng sn xut Taxol v tin cht
Son, S.H.; Choi, S.M.; Lee, Y.H.; Choi,
ca n bng cch nui cy t bo
K.B.; Yun, S.R.; Kim, J.K.; Park, H.J.;
T.cuspidata trn quy m th nghim. Tng
Kwon, O.W.; Noh, E.W.; Seon, J.H.;
sn lng thu c khong 3 mg/l Taxol
Park, Y.G.(2000)
v 74 mg/l taxoid sau 27 ngy nui cy.
Cc dng lp th ca methyl jasmonate v
tc ng ca chng i vi nng sut t
bo v thc y sn xut Taxol v Yukimune, Y.; Hara, Y.; Nomura, E.;
baccatin III c nghin cu trn dch Seto, H.; Yoshida, S. (2000)
treo t bo T.media.
Nui cy t bo T.media gm 2 giai on
c thc hin vi bioreactor 5L c
Cusid, R.M.; Palazn, J.; Bonfill, M.;
khuy trn. Sn lng thu nhn c l
Navia-Osorio, A.; Morales, C.; Piol,
21.12 mg/l Taxol v 56.03 mg/l baccatin
M.T. (2002)
III sau 20 ngy nui cy.
Nhng nh hng ca cht cy
(inoculum) ti kch thc, tui tng
trng ca sinh khi v kh nng sn xut
Taxol trong qu trnh nui cy t bo T.
yunnanensis.

Zang, C.H.; Wu, J.Y.; He, G.Y. (2002)

Dch treo t bo ca T. baccata v T.

wallichiana tng trng trong bioreactor
Navia Osorio, A.; Hermann, G.; Cusid,
20 L, tc tng trng v kh nng tch
R.M.; Palazn, J.; Alfermann, A.W.;
ly Taxol v baccatin III ca 2 loi Taxus
Piol, M.T. (2002)
c ghi nhn.

52

Chng 2: TNG QUAN

Bng 2.2: (Tip theo).
Tm tt nghin cu

Tc gi

Tc ng ng h tr ca cc elicitor
khc nhau c nghin cu trn dch treo
t bo ca T. chinensis, kt qu cho thy
hn hp cc elicitor vi cc tc ng khc Yuan, Y.J; Wei, Z.J.; Miao, Z.Q.; Wu,
nhau c kh nng ng h tr trong khi J.C. (2002)
cc elicitor c tc ng ging nhau li
khng th.
S vn chuyn ca Taxol trong dch treo
t bo T. baccata c nghin cu bng
cch s dng Taxol [14C] c nh du. Fornal, S.; Esposti, D.D.; Navia-Osorio,
Cc phn t c hp th trong c thnh A.; Cusid, R.M.; Palazn, J.; Piol,
t bo (20%) v trong t bo cht (80%), M.T.; Bagni, N. (2002)
cho thy s tch t trong khng bo.
c ch s tch ly Taxol v baccatin III
vi mevinolin v fosmidomycin c
Palazn, J. Cusid, R.M.; Bonfill, M.;
nghin cu trn dch treo t bo T.
Morales, C.; Piol, M.T. (2003)
baccata.
M t cc taxoid ch yu c sn xut
bi dch treo t bo T. media trong iu Ketchum, R.E.B.; Rithner, Ch.D.; Qiu,
kin nui cy bnh thng v vi s b D.; Kim, Y.S.; Williams, R.M.; Croteau,
sung methyl jasmonate.

R.B. (2003)

S nh hng ca cc elicitor trn s sn

xut taxoid v hot ng ca 3-hydroxyBonfill, M.; Palazn, J.; Cusid, R.M.;
3-methylglutaril coenzyme A reductase
Joly, S.; Morales, C.; Piol, M.T. (2003)
trong t bo T. media c nghin cu.

53

Chng 2: TNG QUAN

Bng 2.2: (Tip theo).
Tm tt nghin cu

Tc gi

Nhng nh hng ca sng siu m

cng thp trn cm oxy ha
(oxidative burst), sinh tng hp acid
jasmonic v sn xut Taxol trong dch Wu, J.; Ge, X. (2004).
treo t bo T. chinensis c nghin
cu.
Tc ng lun phin v trc tip ca t
trng ln s tng trng v sn xut

Shang, G.-M.; Wu, J.-Ch.; Yuan, Y.-J.

Taxol trong nui cy t bo T. chinensis
(2004).
var. mairei c nghin cu.
S cm ng ca methyl jasmonate v
salicylic acid (SA) i vi s sinh tng
hp Taxol v cc taxoid c lin quan
trong dch treo t bo ca T. chinensis
Wang, Y.D.; Yuan, Y.J.; Wu, J.C. (2004,
c nghin cu. Kt qu cho thy SA c
2007).
th lm tng qu trnh sn xut Taxol
bng cch ngn chn con ng sinh tng
hp t baccatin III thnh cephalomannine.
Sn xut Taxol v baccatin II bi t bo
T. baccata t do v c nh c nghin
cu trn ba loi bioreactor khc nhau
(khuy trn, sc kh v to sng). Bentebibel, S.; Moyano, E.; Palazn, J.;
Bioreactor khuy trn c hiu qu nht Cusid, R.M.; Bonfill, M.;
trong vic thc y t bo c nh sn Piol, M.T. (2005)
xut Taxol, hm lng Taxol thu c l
43,43 mg/l sau 16 ngy nui cy.

54

Chng 2: TNG QUAN

Bng 2.2: (Tip theo).
Tm tt nghin cu

Tc gi

Phn ng ca dch treo t bo T.

yunnanensis (cy Thng c ngun gc
t tnh Vn Nam, Trung Quc) khi b sc
nhit cng c nghin cu. T bo b
sc nhit 35 - 50oC trong thi gian 30 - Zhang, Ch.; Fevereiro, P.S. (2007)
60ph. Kh nng sn xut Taxol v cm
ng ca t bo trong qu trnh nui cy
gim xung r rt. Vic b sung ABA vo
nui cy cng c kho st.
C ch cm ng sn xut Taxol trong
nui cy t bo T. cuspidate khi b sung
vi Ce4+ hoc methyl jasmonate c
nghin cu. Kt qu cho thy rng s cm Yang, S.; Lu, S.H.; Yuan, Y.J. (2008)
ng lin quan n thnh phn lipid ca
mng t bo. Sn lng Taxol c nng
cao.

Phng php nui cy t bo l mt phng php thay th gp phn vo s

pht trin mi trng bn vng. Phng php ny c mt s u im nh khng
ph thuc vo iu kin thi tit, ma v nhim, v nguyn liu c th pht trin
c lp so vi ngun gc ban u, iu kin c qu trnh v v tr kch
thchy l phng php c tim nng kinh t ln so vi phng php truyn
thng, v n c kh nng sn xut mt lng ln taxoid tinh khit.
Hin nay, Python Biotech l nh sn xut Taxol ln nht bng phng php
nui cy m t bo, s dng h thng fermentor ln ln ti 75000 L. Mt cng ty
khc, Corean Samyang Genex, s dng nui ct t bo Taxus sn xut Taxol
vi tn thng mi l Genexol [27].
2.2.5.3 Bn tng hp t DAB
Taxol cng c sn xut trn quy m ln bng phng php bn tng hp
t baccatin III hoc 10-deacetylbaccatin III, y l mt n lc ln cho vic khm

55

Chng 2: TNG QUAN

ph ra cc phng php ci thin hoc thay th cho qu trnh sinh tng hp t
nhin.
BMS (Bristol-Myers Squibb), l nh cung cp Taxol hng u th gii, c mt
trang tri ln ti 30 triu cy cung cp v v l cho vic tch chit cc hp cht
trung gian. Trong nm 2007, Indena pht trin phng php bn tng hp Taxol
theo mt quy trnh c cp bng sng ch da trn 10-deacetylbaccatin III [27].
S tng hp Taxol v cc hp cht tng t n c bt u vi 10-DAB, hp
cht ny phi c bo v v tr C7 v C10 trc khi acyl ha ti v tr C13. Nu
hp cht 10-acyl, tng t nh Taxol l sn phm mong mun, 10-DAB thng
c bo v bng 7-triethylsilyl ether ca n v sau acyl ha chn lc C10
trc khi c phn ng vi mt chui bn thch hp.
Nhiu cch tip cn khc nhau c s dng ni chui bn vi baccatin III
( c bo v v tr C7 v C10). Phng php da trn -lactam l phng
php c s dng bi BMS bn tng hp Taxol cho n khi n c thay th
bng cc phng php nui cy m thc vt.
-lactam l cht ha hc c pht trin c lp bi Holton v Ojima, y l
mt con ng tng hp Taxol n gin v hiu qu [22].
Cha kha l nonracemic -lactam c th chun b bng cc phng php khc
nhau, nhng tt c u lin quan n phn ng [2+2] cycloaddition (l mt phn
ng pericylic- trng thi chuyn tip ca phn t c hnh dng vng).
Lin kt ca -lactam v baccatin III cho hiu qu v nng sut cao. Lin kt
c thc hin bnh thng vi s c mt ca NaH hoc LiHDMS, v sn phm
t c thng trn 90%.

56

Chng 2: TNG QUAN

Hnh 2.34: Bn tng hp Taxol t DAB [33].

2.2.6 ng dng hp cht Taxol trong iu tr bnh ung th
2.2.6.1 Ung th bung trng
Taxol bc u c ph chun bi FDA (Food v Drug Administration) vo
nm 1992 iu tr cho ph n b ung th biu m bung trng trn c s kt
qu iu tr th nghim truyn dch Taxol lin tc trong 24h. Kt qu ny thc y
cho s m rng iu tr giai on 2.
Trong nghin cu ln u tin, 30% ph n c iu tr trc phn ng
vi kh nng chng ung th. Mt phn ng chnh c xc nh l c th l phn
ng hon ton (s bin mt hon ton bnh, vi cc du hiu bnh thng ca khi
u) hoc l phn ng mt phn (gim tng sn phm trao i cht tt c cc v tr
ca bnh t nht l 50%). Thi gian phn ng trong khong 1 ti 15 thng, trung
bnh l 6 thng. 24 % ph n c biu hin khng li vi iu tr bng liu php
platinum (bnh pht trin tr li trong thi gian 6 thng) cho phn ng vi
Taxol, trong khi t l ny l 40% nhng bnh nhn ti pht sau khi c iu
tr vi plantium (v nhng ngi c th p ng vi iu tr lp li vi liu php
s dng platinum) [19].
57

Chng 2: TNG QUAN

Mc d, qu trnh iu tr lm gim nghim trng lng bch cu trung tnh
trong hu ht cc qu trnh, ngay c khi s dng liu lng thp, nhng thi gian
li tng i ngn v ngi iu tr t b st. Kt qu ca cc th nghim kim
chng cng cho kt qu tng t. Kt qu th nghim ny tt hn ng k so vi
cc phng php ha tr khc cng nh so snh vi kt qu ban u khi iu tr vi
cisplatin.
Trn c s cc kt qu vi liu lng tng thp trn ph n b bnh, Taxol
c kt hp cng vi nhn t kch thch bch cu ht ni vng - l nhn t kch
thch cho php tng liu lng. Phn ng chnh vi kh nng chng ung th xy
l 48% trn cc bnh nhn iu tr liu cao trc . Thi gian ko di s sng l
11.5 thng, tng t nh bo co th nghim trc vi liu lng thp hn.
Thi gian ti pht (relapse-free survival) trung bnh l 6.2 thng, v t l ny l
41% sau 9 thng [19].
Nhng kt qu ny vi liu lng cao hn nu ra kh nng lin h gia liu
lng v s phn ng vi thuc. Cc c im chung ca cc kt qu th nghim
lm sng iu tr cho ph n b ung th bung trng c chng minh trong
mt chng trnh iu tr c tin hnh bi Vin Ung Th Quc Gia (National
Cancer Institute) Hoa K. Thng qua chng trnh ny, Taxol (135 mg/m2 trong
thi gian 24h) c thnh lp bc u cung cp cho ph n ung th bung
trng c tin trin sau khi iu tr vi ba phc . 22% trong 1000 bnh nhn
u tin c phn ng mc d trc h c chun on c nhng c im
khng kh quan [19].
Vi nhn nh rng Taxol v cisplatin c th kt hp an ton, mt bc tip
theo chnh l so snh ch iu tr vi Taxol (135 mg/m2) tip theo l
cisplatin (75mg/m2) vi mt ch iu tr tiu chun vi cyclophosphamide (750
mg/m2) v cisplatin (75mg /m2) trn ph n ung th (khng c iu tr vi giai
on III v IV ca ung th bung trng m c phu thut loi b mt phn
ca khi u). C s phn ng trong nhm cyclophosphamide v nhm Taxol tng
ng l 64 v 77% ph n. Ch iu tr vi Taxol lm gim nguy c ti pht
32% v ko di thi gian sng st trung bnh l 13.8 v 17.9 thng i vi ph n
cc iu tr tng ng vi cyclophosphamide v Taxol [19].
Tuy nhin, thi gian theo di l khng nh gi t l sng st tng th.
Mc d, s gim lng bch cu trung tnh cao hn khi iu tr vi Taxol, nhng
li khng lm tng s nhim khun mu. Nghin cu ny cho thy c s kt hp
58

Chng 2: TNG QUAN

ca Taxol v cisplatin s tr thnh liu php tiu chun mi iu tr ung th bung
trng.
Hai vn quan trng - thi gian iu tr chnh ca Taxol (truyn dch trong
thi gian ngn so vi truyn dch trong thi gian di) v mi quan h gia liu
lng - phn ng trong phm vi liu lng thng thng - ang c nghin cu
ph n ung th bung trng. Nh tho lun, nh hng ca 2 liu Taxol (135
v 175 mg/ m2) v 2 thi gian biu (24h v 3 h/m2) vi phn ng mn cm i vi
tin gy m (premedication) l nh nhau [19].
Thi gian sng st ca nhm liu cao so (175 mg/ m2 ) so vi nhm liu thp
(135 mg/ m2 ) l 19 so vi 14 tun, nhng s sng st ging nhau c 2 nhm
theo c liu lng v thi gian biu. Mc d, liu lng iu chnh c ph
chun l s dng liu 135 mg/m2 trong thi gian 24h trn ph n khng thuc v
c ti pht trong iu tr ung th bung trng, kt qu ny l nhng ng lc
thc y s ph chun iu chnh tip theo l s dng liu chnh l Taxol 175
mg/ m2 trong thi gian 3h.
S tng quan gia liu lng phn ng cng c nh gi trong mt th
nghim lin tc vi ph n khng li iu tr vi platinum l iu tr vi 1 trong
3 liu Taxol 135, 175 v 250 mg/m2 (cng thm kch thch t bch cu ht ni
vng) trong khong thi gian 24h.
Hai th nghim quan trng khc vi ph n b ung th bung trng. Mt th
nghim l ph n c s phu thut ct b mt phn khi u giai on 3 hoc 4 ca
bnh ang c iu tr vi s kt hp ca Taxol v cisplatin hoc dung np ti a
vi liu ch c Taxol hoc cisplatin xc nh xem s hiu qu hn so vi cc
loi thuc ring bit.
Mt th nghim khc l ph n ang giai on 3 ca bnh c iu tr
bng cch ct b mt phn khi u ti u c iu tr vi Taxol v cisplatin,
cyclophosphamide v cisplatin, hoc Taxol v iu tr chuyn su vi cisplatin v
carboplatin. Liu cyclophosphamide v cisplatin c th c ngng tc ng ca
thuc trn bnh sau phu thut l thp hn so vi Taxol v cisplatin, nghin cu
ny s cung cp mt s kt qu nh gi tc ng ca Taxol trn mt s nhm
bnh nhn c d on tin trin trong iu tr.

59

Chng 2: TNG QUAN

2.2.6.2 Ung th v
Hot tnh chng ung th quan trng trn ph n b ung th v di cn c
chng minh vi Taxol trong thi gian 24h.
Trong s 25 ph n nhn c khng nhiu hn 1 phc ha tr cho bnh di
cn, 50% c phn ng v thi gian trung bnh bnh c tin trin l trong thi
gian 9 thng. Mt th nghim khng nh trong Taxol (250 mg/m2) c
a ra vi cc kch thch t bch cu ht ni vng cho cc ph n (nhng ngi
nhn c s iu tr b sung hoc khng c iu tr trc ) khng nh
cho hot ng ny. Kt qu l 62 % bnh nhn c phn ng [19].
Trong c 2 th nghim, phn ng xy ra tt ca cc v tr di cn ca bnh ung
th (kh iu tr vi anthracycline). Nhng kt qu ng kch l vi Taxol trn ph
n ung th v c so snh vi kt qu c bo co trong cc nghin cu ban
u vi anthracycline, mt trong nhng tc nhn c hiu qu trong iu tr ung th
v. nh gi tm quan trng chung ca cc kt qu ny v d on vai tr cui
cng ca Taxol trong ung th v lin quan ti s phn ng ca khi u l kh khn,
tuy nhin, khi nhng nghin cu ban u c khng nh trn nhng ph n t
c iu tr trc .
Tip theo cc nghin cu trn nhng ph n ( c iu tr trc ) khng
nh nhng kt qu ban u, mc d kh nng phn ng thp hn. Kh nng phn
ng dao ng trong khong 38% ph n ( c iu tr trc vi mt phc
cho bnh di cn) ti 17% (trn nhng ngi trc y c iu tr vi ba
hoc nhiu phc cho bnh di cn). T l phn ng tng th trn nhng ph n
rt nhy cm vi hoc c biu hin khng vi anthracycline l nh nhau. S thiu
hon chnh trong khng cho (cross-resistance) gia Taxol v anthracycline cho
thy s kt hp to thnh thuc hn hp (multidrug) c th khng quan trng.
S pht trin xa hn na ca Taxol iu tr cho ph n ung thu v s lin
quan n vai tr ca n trong cc giai on u v giai on cui cng trong iu
tr h tr. Trong mt lot cc th nghim lm sng ca Eastern Cooperative
Oncology Group iu tr cho nhng ph n b bnh ung thu v di cn (trc y
khng c iu tr vi Taxol hoc doxorubicin hoc c hai). Nu liu php da
trn Taxol chng minh s vt tri th n s c kt hp vo trong th nghim
h tr. Ha tr liu cng cao lin tc vi doxorubicin tip sau l Taxol v
cui cng l cyclophosphamide ang c nh gi trong iu tr h tr cho bnh
60

Chng 2: TNG QUAN

nhn c nguy c cao sau khi iu tr dt im cc khi u chnh bng phu thut v
x tr.
c tnh hu ch ca Taxol nh l mt phn ca iu tr h tr i vi bnh
nhn c nguy c cao cng ang c nghin cu trong mt th nghim lm sng
quy m ln vi cng cao, trung bnh v thp vi doxorubicin v
cyclophosphamide tip l Taxol v cui cng l tamoxifen hoc iu tr ch c
tamoxifen.
Thi gian v liu lng ti u ca Taxol iu tr cho ph n ung th v di
cn ang c nh gi. Theo mt kt qu nghin cu trc l truyn dch
Taxol trong thi gian 3h 135 mg/ m2 v 175 mg/ m2 cho thy khng c s khc
bit ng k trong t l phn ng (29% [liu cao] v 22% [liu thp] v thi gian
ko di s sng trung bnh l 11.7 thng [liu cao] v 10.5 thng [liu thp]. Cng
khng c s khc bit t l gia ph n c s khng vi iu tr bng
anthracycline v khng c s khng thuc. Cc kt qu ca th nghim ny cng
nh trc giai on 2 ca th nghim, dn ti ph chun quyt nh s dng Taxol
liu lng l 175 mg/ m2 trong thi gian 3 gi, iu tr ung th v di cn sau
khi ha tr liu kt hp tht bi hoc ti pht trong vng su thng vi ha tr liu
b sung [19].
Mt th nghim vi quy m ln M cho ph n tri qua mt phc ha
tr liu ung th v hoc ti pht sau khi iu tr vi liu ti u, vi cc liu lng
l 175, 210 v 250 mg/m2. Ti u thi gian iu tr cng ang c th nghim
trn nhng ph n (khng s dng ha tr liu hoc ang c iu tr) vi thi
gian 3h hoc 24h truyn dch Taxol (250 mg/m2) vi kch thch t bch cu ht ni
vng [19].
2.2.6.3 Ung th phi
Taxol cng c nh gi trn cc bnh nhn ung th phi (nonsmall-cell
lung cancer) cha c iu tr, vi liu lng bt u (200 ti 250 mg/m2) vi
thi gian 24h.
Trong mt nghin cu, t l phn ng l 24%, thi gian phn ng trung bnh l
27 tun v thi gian sng trung bnh l 40 tun (56 tun cho nhng ngi c phn
ng vi iu tr). Trong mt nghin cu ngu nhin giai on 2 cng bao gm
cc nhn t nghin cu l piroxantrone v merbarone, t l phn ng l 21, 2.3 v
0% cho Taxol, piroxantrone v merbarone, tng ng vi tng loi. Thi gian
61

Chng 2: TNG QUAN

trung bnh cho phn ng vi Taxol l 6.5 thng, t l sng 1 nm l 41.7 %, thi
gian tn ti trung bnh l 24.1 tun [19].
Trong 2 nghin cu khc, t l phn ng nhng bnh nhn ung th phi
(khng c iu tr trc ) vi Taxol (250 mg/ m2) c v khng c nhn t
kch thch bch cu ht ni vng l 41 v 34 % [19]. Kt qu c so snh vi
mt s tc nhn c lp khc s dng chng li cc khi u ny.
Cc th nghim tip theo trong ung th phi s nh gi hiu qu ca s kt
hp ha tr liu. Hai phc iu tr tiu chun c so snh trn cc bnh nhn
ung th phi (nonsmall-cell lung cancer) cha c iu tr trc . Mt phc
bao gm etoposide (100 mg/ m2) vo ngy 1-3 v cisplatin (75 mg/m2), mt
phc iu tr khc bao gm cisplatin (75 mg/m2) v truyn dch Taxol trong 24h
liu thp (135 mg/m2) v liu cao (250 mg/m2) vi nhn t kch thch bch cu
ht ni vng.
2.2.6.4 Ung th u v c
Taxol (250 mg/m2 trong 24h vi cc yu t kch thch bch cu ht ni vng)
chng minh hot ng trn bnh nhn ung th di cn t bo biu m u v
c (khng c iu tr trc ). Trong mt th nghim quy m ln vi 2 giai
on, 43% bnh nhn c phn ng, mt kt qu kh quan khi so snh vi t l ca
cc nhn t c lp thng thng khc [19].
Vai tr ca cc liu lng khc nhau cng nh hiu qu ca thuc khi kt hp
vi cc loi thuc khc nh cisplatin, fluorouracil, v ifosfamide, ang c
nghin cu.
2.2.6.5 Cc loi khi u khc
Mc d tc ng ca Taxol trong khi u c tnh trong nghin cu tin lm sng
v mt s bnh nhn c phn ng trong giai on 1 ca th nghim, t l phn
ng ch l 12 v 18 % trong giai on 2 ca nghin cu [19].
Giai on 2 th nghim ung th tuyn tin lit, ung th i trc trng, ung
th thn, ung th tuyn ty v ung th d dy khng c kt qu. T l p ng thp
ch l 17% i vi bnh nhn ti pht vi non-Hodgkins lymphoma [19].
Trong mt nghin cu khc, 12 v 29% bnh nhn b bnh mn tnh s cp v
ti pht vi non-Hodgkins lymphoma c phn ng vi Taxol [19].
62

Chng 2: TNG QUAN

Mt khc, Taxol (250 mg/m2) iu tr trong 24h vi nhn t kch thch bch cu
ht ni vng c tc ng c li trn bnh nhn ung th tinh hon, ung th bng
quang v ung th thc qun. 42% bnh nhn khng c iu tr vi ung th thc
qun (advanced transitional-cell carcinoma of the bladder) c phn ng. T l
phn ng trn bnh nhn b ung th thc qun (advanced esophageal cancer) l
32%, mc d t l ny thp hn trong cc bnh ung th ng tiu ha khc, phn
ng xy ra vi c adenocarcinoma (ung th biu m bt ngun t cc m tuyn,
34%) v squamous-cell carcinoma (ung th biu m vy, 28%) [19].
Ngoi ra, 24% bnh nhn khng vi cisplatin ung trong iu tr ung th t bo
mm (germ-cell cancer) c phn ng vi Taxol trong mt nghin cu khc [19].

63

Chng 3: MT S KT QU VI NHN GING TAXUS

Chng 3: MT S KT QU VI NHN GING TAXUS

3.1 Vi nhn ging Taxus baccata L.
Phng php vi nhn ging c la chn nhn ging T. baccata c pht
trin trong cng mt thi gian, tuy nhin s lng kt qu cng b cn hn ch
(Taxus mairei Chee, 1995; Chang v cng s, 1998, 2001; Taxus baccata
Majada v cng s, 2000) ( theo Ewald D., 2007) [17].
S to phi soma ca Taxus, ch yu t mu cy cn non cng nh phi hp
t cha trng thnh cng c bo co, nhng s ti sinh ca thc vt qua qu
trnh hnh thnh phi v s pht trin ng b ca cy kh xy ra. (Taxus
brevifolia Ewald v cng s, 1995 v Taxus chinensis Qiu v cng s, 1998)
[17]. Ngoi ra, phi soma c t l to thnh chi mi rt thp.
Cc th nghim v nhn ging v tnh Taxus trc y khng thnh cng so s
dng nguyn liu ban u t cy trng thnh. V vy, mt phng php vi nhn
ging khc c la chn nhn ging v tnh dng Taxus c chn.
3.1.1 Mi trng nui cy
Mi trng nui cy cho qu trnh vi nhn ging c lit k trong bng 3.1.
Nng ca cc thnh phn c bn (pha long hoc tng nng ) t cc mi
trng thng thng hin c, trong khi cc cht iu ha sinh trng, b sung
c cp trong tng phn ring bit.
Da trn cc th nghim trc , mi trng Woody Plant Medium (WPM,
theo Lloyd v McCown, 1981) c kh nng h tr cho qu trnh tng trng v sc
sng ca mu cy Taxus tt nht. Do , qu trnh to chi cng nh ko di chi
u s dng mi trng ny.
Mi trng thch hp cho s pht sinh r l mi trng LS bin i (L9, da
trn LS theo Linsmaier v Skoog, 1965).

64

Chng 3: MT S KT QU VI NHN GING TAXUS

Bng 3.1: Thnh phn mi trng dinh dng c s dng cho qu trnh vi
nhn ging [16].
Nguyn t a

Nguyn t vi

Ngun Carbon

lng

lng

(g/l)

20 sucrose

5.7

W double

20 sucrose

5.7

L9

1/3

5 sucrose

5.7

Mi trng

pH

(Nguyn t a lng, nguyn t vi lng c thay i t l so vi mi trng

dinh dng c bn ban u).
3.1.2 Cc giai on ca qu trnh vi nhn ging
3.1.2.1 Chun b mu cy
a. iu kin sinh trng ca cy m
Cnh ca cy trng thnh c la chn v thu hoch trong thng 9, c
kch thch ra r vi hn hp c cha 2 g/l indole-3-butyric acid (IBA) v c
phun sng di p lc cao (high pressure fog) trong nh knh (hnh 3.1 A). Cc
cy bn r c s dng lm nguyn liu nui cy. Chi bn v nh chi
ngn c thu hoch lm mu cy sau khi x l s b vi thuc dit nm (0.2%
Euparen by Bayer, 50% dichlorfluanide) trong 24h [17].
b. Kh trng mu cy
B mt ca mu cy c gi kh cho php kh trng c hiu qu. Do ,
khng c ra trc khi kh trng v cy khng c ti nc 1 ngy trc khi
s dng.
Kh trng c thc hin bng cch ra mu cy vi dung dch thy ngn
cloride (0.25%) vi 1 git Tween 80 t 10-15 ph. Sau , mu cy c ra sch
ba ln vi nc v trng v t trn mi trng dinh dng trong bnh erlen 100
ml.
Mc tiu ca giai on ny l thu c mt lng ln mu cy v trng m
vn cn kh nng tng trng.

65

Chng 3: MT S KT QU VI NHN GING TAXUS

Hnh 3.1: Vi nhn ging T. baccata [17].

A. Cnh gim. B. Nhn ging chi. C. S ko di ca chi. D. R hnh thnh trn than
bn. E. L sp xp i xng ta trn [bn tri] hoc hnh ch V [bn phi]. F. Cy sau
khi chuyn ra t.

66

Chng 3: MT S KT QU VI NHN GING TAXUS

3.1.2.2 S pht sinh v duy tr ca chi ngn
a. Hnh thnh chi bn v qu trnh nhn ging
Th nghim c hiu qu i vi cytokinin, v d nh benzylaminopurine (BAP)
kch thch chi bn v chi ngn pht trin trn mi trng dinh dng c b sung
than hot tnh.
Cc th nghim trn cc mi trng c bn khc nhau cho thy Woody Plant
Medium (W) l mi trng c hiu qu nht cho qu trnh tng trng v kh nng
sng st ca mu cy. Mi trng Woody Plant Medium vi 1.5 mg/l zeatin (c
gi l Wz) c s dng cho cm ng to chi bn. i khi, zeatin c thay th
bi 2iP (N6-(2-isopentenyl) adenine-2.84 mg/l).
Mu cy c gi di iu kin nh sng lin tc (30 E m2s1, 650 nm)
nhit 23oC. Mi trng Wz lm gam s ko di ca chi, v hnh thnh chi
bn (Hnh 3.1 B). Nu nng mi trng c bn tng gp i (W-double) v s
dng nng zeatin vi nng tng t nh mi trng Wz (Wz-double) c th
ci thin ng k s ko di ca chi ngn, s lng chi bn bn v chiu di l.
Tuy nhin, s dng lin tc Wz-double s gim t l nhn ging.
Mi trng Wz c s dng do c tc ng tch cc ln s tng sinh chi
nch vi t l 1.2 - 1.8 chi/ mu cy/ thng. Vic b sung 200 mg/l spermidine c
tc ng tch cc ln s to chi [17].
Kt qu l thu c chi c kh nng to r trn mi trng ny.
Vn hay gp phi trong qu trnh nhn ging l s c mt ca vi khun
endophytic, do cn b sung 500 mg/l ticarcilline gii quyt vn ny.
b. Ko di chi
Ko di ca chi bn
Mt lot cc th nghim lin quan n qu trnh nhn ging ca on thn c
mang chi bn bng cch so snh cc giai on ko di ca ca chi vi h thng
s dng cho cy thng rng l, cc th nghim ny khng thnh cng v khng th
kch thch s pht trin ca chi bn trong cc on thn.
Mt khc, s ko di ca nh chi ngn trong th nghim ny trn mi trng
(W-double) kt hp vi zeatin v kinetin (0.66 + 0.64 mg/l) vi vic b sung PVP
v arginine (100 mg/l mi loi) c xem l tt nht. S ko di ca chi bn
67

Chng 3: MT S KT QU VI NHN GING TAXUS

khng xy ra trn mi trng ny nhng nu chi c cy chuyn sang mi
trng khng cha phytohormone ri tip theo chuyn sang mi trng c
thidiazuron nng thp (TDZ, 0.01 mg/l) th s ko di chi xy ra.
Ko di chi
Chi ngn ko di nht khi s dng mi trng W - double c b sung 0.01
mg/l TDZ kt hp vi 200 mg/l spermidine. Bng cch ny c th kch thch s
ko di ca chi t cm chi nch l (hnh 3.1 C). S kt hp ny cng thc y
s pht trin ca l trn chi (chiu di l). Chi ban u c chiu di 15 mm, qua
2 ln cy chuyn (8 tun), chiu di chi tng ln n 35 40 mm [17].
Nng cao TDZ c ch s pht trin ca l v s ko di ca chi.
T th nghim to r c th kt lun rng s to r c ci thin khi chi c
nui trong mi trng c cha spermidine trc khi chuyn sang mi trng to
r. V vy, chi sau khi hnh thnh c cy chuyn trn mi trng khng c
hormone 4 tun, tip theo chuyn sang mi trng c cha spermidine trc khi
cm ng to r.
3.1.2.3 S ra r in vitro
Tt c cc bc cm ng ra r v pht trin c thc hin 15 - 17 oC v chu
k chiu sng l 16h (nh sng trng c c cng bc x 30 E m2s1).
Mi trng nui cy trc khi to r cng nh hng ti kt qu ca qu trnh
ra r sau ny. Zeatin c trong mi trng c s dng trc khi ra r lm gim t
l r to thnh. Spermidine kt hp vi TDZ (0.01 mg/l) lm tng t l to r cng
nh sc sng ca chi. So snh vi cc loi cy l kim khc, s to r ca cy
Thng i hi thi gian ko di n 7 thng [17].
a. S cm ng s to r vi auxin
on chi (chiu di 30 - 40 mm) c cm ng trn mi trng L9 c cha 2
mg/l IBA trong 2 tun [17]. IBA c tc ng hiu qu hn khi c so snh vi
NAA (naphthalene acetic acid). Chi c ct 1 phn (2 mm) trc khi t chng
trn mi trng cm ng.

68

Chng 3: MT S KT QU VI NHN GING TAXUS

b. S cm ng s to r vi Agrobacterium rhizogenes
Chi c chun b bng cch tng t nh cm ng vi auxin sao cho gc ca
chi c th tip xc vi mi trng, qu trnh c thc hin trong 24h vi dung
dch c cha nhiu chng Agrobacterium rhizogenes (OD = 0.6 600 nm - mi
trng dinh dng 20E; Werner v cng s, 1975) c b sung 19.6 mg/l
acetosyringone.
S hnh thnh r v t l sng st trn mi trng cm ng bi chng A.r.
Marburg l tt nht khi so snh vi s dng IBA. M so hnh thnh cng nh s
phn hy ca gc chi gim trong qu trnh di pht trin ca r sau khi cm
ng bi Agrobacterium rhizogenes v c bit l chng A.r. Marburg. S a dng
ca ca chng Agrobacterium c lin quan n trng thi chung ca chi trong qu
trnh to r [17].
Do , nhiu chng Agrobacterium rhizogenes c s dng cm ng to
r, bi v cc cht cm ng t nhin c kh nng h tr cho qu trnh pht trin ca
r, trong mt s trng hp kt qu tt hn so vi cm ng auxin.
c. S pht trin ca r
Sau thi gian cm ng, chi c t trc tip trong nhng ming than bn
bo ha nc. Nhng ming than bn c t vo trong nhnh hp knh nh.
Trng ru gia cc ming than bn ngn cn s tn cng ca nm.
Thi gian r pht trin ko di t 5-7 thng cho n khi r c th thy c bn
ngoi ming than bn (hnh 3.1 D). V l do ny, hm lng m ca ming than
bn phi c kim tra hng tun ( trnh lm kh).
3.1.2.4 Trng cy trn rung
Sau khi r hnh thnh, c chuyn vo hp hun luyn r (RONAASH Ltd.
Scotlv, 4.5 4.5 cm,cao 20 cm, mi ci c khong 40 cy con) c bn phn
(EINHEITSERDE, mu VM) v phun sng p lc cao ( m khng kh 95%).
m khng kh c gim dn trong thi gian 1 thng.
Ging nh cc cy l kim khc, c 2 trng thi khc nhau c th quan st trong
qu trnh tng trng ca cy khi chuyn ra t, tng trng hng thng v
hng nghing. i vi s tng trng ca Taxus, trng thi ny c lin quan cht
ch n cch sp xp l trn chi ko di.
69

Chng 3: MT S KT QU VI NHN GING TAXUS

L i xng ta trn l mt ch s cho tng trng hng thng, tng t nh
cy ging bnh thng. L xp thnh hai hng, l mt ch s tng trng ging
cnh (branch-like) hoc tng trng hng nghing (Hnh 3.1 E).
Hai c im ca s tng trng c quan st trn chi trong mi trng nhn
ging in vitro mt s dng v tnh. Chi nh (<20 mm) ch yu l i xng ta
trn (t l chi l 10:1), trong khi chi di hn (>20mm) cho thy t l ny gn nh
bng nhau (1:1). S lng chi hnh thnh l hnh ch V tng ln trong sut thi
gian ko di. C mt s lin quan cht ch ca qu trnh tng trng (hng thng
hoc hng nghing) v cch sp xp l trn chi (ta trn hoc hnh ch V).
Chiu cao sau khi chuyn ra t(>40 mm), chi vi l i xng ta trn thng
c hnh thnh trn nh chi ca l sp xp hnh ch V. Tuy nhin, mi quan h
ca chi vi cch sp xp l trn chi l 1:1 [17].
Quan st trong thi gian tip theo, mt s cy biu hin s tng trng hng
thng s th hin s tr ha (rejuvenation). Kim tra r ct t cy trng thnh dn
ti kt lun rng tng trng hng thng ca cy vn cha t c kch thc
chun sau thi gian 6 nm c trng trn t. iu ny c th l do s khc bit
sinh l ca m Taxus mu cy ban u t nhn ging bng gim cnh.
3.1.3 Kt lun
Vi nhn ging Taxus baccata t cnh gim l c th thc hin c. Cm ng
ca chi bn l bc cn thit u tin bc tip theo l s ko di ca chi v
cm ng to r. S cm ng ko di ca r c din ra trn mi trng gn ging
nh t. Chng Agrobacterium rhizogenes c th ci thin s hnh thnh r v t l
sng ca cy.
3.2 Vi nhn ging Taxus wallichiana Zucc.
Cho n nay, hai loi Taxus c xc nh Vit Nam l T. chinensis v T.
wallichiana. T. chinensis phn b cc tnh pha Bc, trong khi T. wallichiana
mc trn ni granite cao 1500m ca Lm ng.
Ch c mt vi nghin cu v s ti sinh ca T. wallichiana c bo co.
K thut gim cnh c p dng trong thi gian 1994 - 1996 bo tn T.
wallichiana Phn Vin Sinh Hc Lt, vi t l r hnh thnh sau 90 ngy l
38% (hnh 3.2 A), chiu di r trung bnh l 6 - 8 cm sau 3 thng [15].

70

Chng 3: MT S KT QU VI NHN GING TAXUS

i vi cng tc bo tn v nhn ging ca T. wallichiana, mt loi cy c
nguy c tuyt chng cao v c gi tr, phng php nhn ging in vitro c pht
trin thng qua cm ng chi, ti sinh chi v r.

Hnh 3.2: K thut gim cnh c p dng cho Taxus wallichiana [15].
A. S ra r ca cnh. B. Cnh gim c ra r sau 3 thng.
3.2.1 Mi trng nui cy
Mi trng MS c cha khong v vitamin c b sung vi 20 g/l sucrose
c s dng lm mi trng m trong sut qu trnh th nghim.
Auxin NAA, IAA v IBA, cytokinins BA c c b sung vo mi trng
nui cy vi cc nng khc nhau (bng 3.2, 3.3 v 3.4).
Than hot tnh cng c b sung gim tc ng ca hp cht phenolic
c to ra t mu cy (Pan v Staden, 1998) (theo D.T Nhut, 2007) [15]. Mi
trng c lm c vi 9g/l agar.
Gi tr pH ca mi trng c iu chnh 5.8 - 5.9 bng cch thm vo
dung dch NaOH 1N v KCl 1N trc khi kh trng 121oC, 1 atm (1.02

10

Pa).

71

Chng 3: MT S KT QU VI NHN GING TAXUS

Bng 3.2: Nng cc cht iu ha sinh trng v than hot tnh b sung vo mi
trng MS c s dng cho qu trnh tng trng v ko di ca chi [15].
Than hot tnh

Mi trng

NAA (mg/l)

IAA (mg/l)

IBA (mg/l)

C1

C2

C3

C4

C5

C6

C7

C8

C9

C10

2.5

C11

2.5

(g/l)

Bng 3.3: Nng ca cht iu ha sinh trng BA v than hot tnh c b sung
vo mi trng MS cm ng s chi bt nh [15].
Mi trng

Mi trng c
bn

BA (mg/l)

Than hot tnh

(g/l)

AB1

MS

AB2

MS

AB3

MS

AB4

MS

72

Chng 3: MT S KT QU VI NHN GING TAXUS

Bng 3.4: Nng cht iu ha sinh trng, vitamin B1 v than hot tnh b sung
vo mi trng MS v mi trng MS cm ng s hnh thnh r [15].
Mi

Mi trng

NAA

IAA

IBA

Vitamin

Than hot

trng

c bn

(mg/l)

(mg/l)

(mg/l)

B1 (mg/l)

tnh (g/l)

R1

MS

10

R2

MS

10

R3

MS

10

R4

MS

10

R5

MS

10

R6

MS

10

R7

MS

10

R8

MS

10

R9

MS

10

R10

MS

2.5

R11

MS

2.5

R12

MS

2.5

R13

MS

2.5

iu kin nui cy
Mi trng nui cy trong phng th nghim 25 2C, m tng i 7580%, cng chiu sng t 4550 mol.m2.s1, chu k chiu sng l 16h.
Mu cy s dng l thn non di t 1.5-2 cm v/hoc chi di 2-2.5 cm.
3.2.2 Cc giai on ca qu trnh vi nhn ging
3.2.2.1 Chun b mu cy
Mu cy c la chn t cc cnh cn non khng c mang nhiu chi (hnh
3.3). Sau 15 n 25 ngy, thu mu cy, la chn nhng mm cn non, mu xanh,
khe mnh.

73

Chng 3: MT S KT QU VI NHN GING TAXUS

Hnh 3.3: Thu thp mu cy T. wallichiana [15].

A. T. wallichiana 5 nm tui Phn Vin Sinh Hc Lt. B. Cnh c mang chi
ng. C. Chi. D. Chi non.
3.2.2.2 Kh trng mu cy
Sau khi thu thp, cnh, thn non v chi c ct thnh nhng on di 5 cm
ra nhiu ln bng nc ct v trng. Nhng mu cy trong cht ty long t 2530 ph, v ra sch di vi nc chy t 1.5 n 2h.
Kh trng b mt vi ethanol 70% trong 30s, 1/1000 HgCl2, 1 n 2 git
Tween-80 0.01% trong thi gian 10-12ph. Sau , ra sch bn ln vi nc ct
v trng.

74

Chng 3: MT S KT QU VI NHN GING TAXUS

Bng 3.5: T l nhim ph thuc vo loi mu cy [15].
Mu cy

T l nhim

Cnh 45 ngy tui

100%

Thn non 25 ngy tui

v chi 14 ngy tui

10%

M t
Mt s mu cy b nhim, ha nu v
cht.
Mu cy khng b nhim, sng v
tng trng nhanh chng.

Mu cy l tng nht thu thp v kh trng l cnh non 25 ngy tui v

chi 14 ngy tui.
Sau khi kh trng, mu c ct thnh on di 2 - 2.5 cm, v t dc trn mi
trng nui cy.
3.2.2.3 Cm ng chi
Hu ht l ca mu cy t thn c mu xanh v cho thy s pht trin bnh
thng, trong khi l ca mu cy t chi c mu vng, sau chuyn thnh

Mu cy c s tng trng ca chi (%)

mu nu v cht sau 8 tun nui cy.

Mi trng

Hnh 3.4: T l mu cy T. wallichiana trn cc mi trng MS khc nhau

(khng c s c mt ca than hot tnh) cho thy s ko di ca chi bn [15].
75

Chiu di chi (cm)

Chng 3: MT S KT QU VI NHN GING TAXUS

Mi trng

Hnh 3.5: Chiu di chi trung bnh (cm) ca mu cy T. wallichiana trn mi

trng MS sa i khc nhau (khng c than hot tnh)[15].
Mi trng C4 l tt nht cho s ko di ca chi, trong khi mi trng C7
thch hp cho s pht trin ca chi bn.
Nng auxin cao c ch s pht trin ca ca chi hoc chi nch v dn ti
s hnh thnh m so. Chi ko di tt hn trn mi trng c cha nng thp
cht iu ha tng trng (PRGs) (Chang v cng s, 2001) [12]. Hu ht mu cy
nng auxin 3 mg/l hoc 5 mg/l cho thy tc tng trng chm hn, sn
xut hp cht phenolic v to thnh m so.
khc phc tnh trng ny, c th b sung thm than hot tnh (2 g/l) vo mi
trng nui cy gim s hnh thnh hp cht phenolic v thc y s ko di
ca chi.
Nh vy, mi trng tt nht cm ng chi l mi trng c 1 mg/l auxin
c b sung thm 2g/l than hot tnh.
Chi mi c tit trng v tng trng trong iu kin in vitro. Sau , chi
t nguyn liu ban u (ex vitro) v chi pht trin trong iu kin in vitro sau 7
tun tui c cy vo mi trng nui cy.

76

Chng 3: MT S KT QU VI NHN GING TAXUS

Sau 10 tun nui cy, tt c mu cy ex vitro tng trng rt tt, l c mu
xanh, trong khi mu cy t chi in vitro 7 tun tui khng c s tng trng v
l c mu vng.
Bng 3.6: nh hng ca chi v thn c ngun gc in vitro v ex vitro trn
chiu di ca chi bn (cm) v t l mu cy c mang chi [15].
Mi
trng

Mu cy

Chiu di ca

Mu cy c chi

chi bn (cm)

(%)

Ghi ch

C10

Thn e

3.00 0.15

100

L xanh

C11

Thn e

6.25 1.25

85.7

L xanh

C10

Thn i

1.93 0.92

52.9

L xanh

C11

Thn i

2.29 1.04

56.3

L xanh, 15.6 %

C10

Chi e

4.24 1.69*
2.21 1.59

100

36.8% c chi bn v 52.6

% c l vng

C11

Chi e

3.92 1.69*
3.75 2.25

85.7

Chi di nht l 9.5 cm.

28.6 % c l vng, theo sau
s tng trng ca chi bn.

C10

Chi i

Khng c s tng trng.

L ha nu v cht

C11

Chi i

Khng c s tng trng.

L ha nu v cht

Ghi ch:
Thn v chi e: ngun gc thn t ex vitro (thn v chi mi c kh trng).
Thn v chi i: thn v chi in vitro.
*: chiu di nh chi.
t cnh non t nguyn liu ban u, di 1.5 - 2cm trn mi trng nui cy.
Theo cc nghin cu trc , cytokinin cn thit cho cm ng to chi trong cy
l kim v BA l cho hiu qu tt nht i vi Taxus mairei (Chang v cng s,
2001) [12].
Cm ng v nhn chi bt nh t cnh non cho hiu qu trn mi trng c
nng thp cytokinin (1 mg/l). 22.4 % mu cy hnh thnh chi bt nh trn mi
trng sau 6 tun nui cy, vi chiu di trung bnh l 0.75 cm. T l ny tng ln
87.5% sau 12 tun nui cy [15].
77

Chng 3: MT S KT QU VI NHN GING TAXUS

Trong khi cng mi trng ny (1 mg/l BA) c b sung than hot tnh
(2g/l), t l mu cy to chi bt nh ch l 18% nhng chiu di trung bnh l 1.3
cm. Sau 12 tun nui cy, c 41.9% mu cy to chi bt nh, 54.2% mu cy
hnh thnh m so [15].

Hnh 3.6: Vi nhn ging in vitro T. wallichiana [15]. A. Chi ti sinh trong
mi trng c cha 1 mg/l IAA. B1. Chi ko di trn mi trng c cha 1 mg/l
IAA. B2. Cy non. C1 v C2. Cm ng chi trn mi trng b sung 1 mg/l BA.
Nh vy, khi c mt than hot tnh chi tng s ko di nhng s lng chi
gim. Webb v cng s (1988) cng chng minh rng s ko di ca chi c
kch thch bi than hot tnh nhng than hot tnh cng c ch cm ng chi khi n
c c BA (theo D.T Nhut, 2007) [15]. iu ny gii thch ti sao t l cm ng

78

Chng 3: MT S KT QU VI NHN GING TAXUS

chi trn mi trng khng c than hot tnh li cao hn hn so vi mi trng c
than hot tnh sau 12 tun nui cy.
Nh vy, mu cy T. wallichiana, cm ng chi mi trng c BA b sung
hoc khng b sung than hot tnh. Mi trng khng c than hot tnh s lng
mu cy nhiu hn nhng chiu di chi li ngn hn so vi mi trng c b
sung than hot tnh.
3.2.2.4 S to r in vi tro
Chn v ct chi (di 1.5 - 2 cm) cm ng to r v t trn mi trng nui
cy c b sung auxin vi cc nng khc nhau.
Sau 10 tun nui cy, mi trng c hoc khng c than hot tnh u cho thy
s tng trng mnh m, l c mu xanh v khng c s hnh thnh m so. Tuy
nhin, s hnh thnh r ch xy ra trn mi trng c b sung IBA (5 mg/l) vi t
l l 1-2 r/ mu cy cho thy IBA hiu qu hn so vi NAA.
3.2.3 Kt lun
Cm ng to chi, ko di chi v ti sinh r m ra mt kh nng mi cho nui
cy t bo T. wallichiana quy m ln c s dng cho khai thc Taxol v 10deacetyl baccatin III.
Cng nh vi nhn ging Taxus baccata, vi nhn ging T. wallichiana c th
thc hin c t cnh gim. Thng qua phng php vi nhn ging in vitro c
th sn xut mt s lng ln cy trng phc v cho cng tc bo tn v nhn
ging T. wallichiana.

79

Chng 4: KT LUN

Chng 4: KT LUN
Cy Thng l mt loi dc liu qu c ng dng nhiu trong y hc, c
bit l trong iu tr ung th. Thng c phm vi phn b rng trn th gii tri
di t vng n i n vng cn nhit i, nhng phm vi ny ngy cng b thu
hp do tnh trng khai thc qu mc, cng nh iu kin sinh trng kh khn ca
loi. Mt s loi l loi c hu t nhin phn b trong mt khu vc nht nh nh
Taxus floridana, Taxus globosa. Hin ti, hu nh tt c cc loi Thng ch
yu c dng lm cy cnh, ch mt s loi nh Taxus brevifolia c trng theo
quy m ln lm nguyn liu trong dc phm.
Vit Nam xc nh c hai loi l Thng Bc v Thng Nam, qun
th c pht hin ca hai loi ny nh, phn b ri rc. y l loi c nguy c
tuyt chng cao cn c nhng bin php bo tn cng nh nhn ging loi thc vt
c gi tr ny.
Taxol l hp cht chnh trong cy Thng , tnh cht ha hc, cu to, c ch
tc ng i vi bnh ung th cng nh con ng sinh tng hp v ang c
nghin cu. Taxol c ph chun trong iu tr ung th bung trng, ung th
v di cn, Taxol cng c tin hnh th nghim lm sng trong iu tr ung th
phi, ung th u v cHp cht Taxol c th c thu nhn bng nhiu phng
php nh tch chit t v v l cy Thng , tng hp bng phng php ha
hc, bn tng hp t DAB, hay gn y nht l thu nhn bng phng php nui
cy t bo. Phng php nui cy t bo thu nhn hp cht Taxol c th thay
th cc phng php khc, gp phn pht trin bn vng mi trng t nhin, m
rng quy m sn xut ln p ng c nhu cu iu tr ung th ngy cng cao.
Vi nhn ging Thng c thc hin thnh cng trn Taxus baccata,
Taxus wallichiana m ra mt bc pht trin quan trng trong cng tc bo tn v
nhn ging Thng , c bit l Vit Nam.

80

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