B Y T

TRNG I HC DC H NI

TRNG MINH PHNG

NGHIN CU THNH PHN HA

HC V TC DNG GIM LO LNG,

CI THIN TR NH CA MN KINH

T ( VITEX TRIFOLIA L. )

KHA LUN TT NGHIP DC S

Ngi hng dn:

1. TS. Phm Nguyt Hng

2. ThS. Phm Thi H Vn

Ni thc hin:

1. Vin dc liu Vit Nam

2. B mn Dc hc c truyn

trng i hc Dc H Ni

H NI 2014

MC LC

T VN ...................................... ................................................................. 1

Chng 1: Tng quan .......................................................................................... 2

1.1. Thng tin chung ............................................................................................. 2

1.1.1. V tr phn loi ca chi Vitex ....................................................................... 2

1.1.2. c im chung ca chi Vitex ..................................................................... 2

1.1.3. c im ca loi Vitex trifolia L.. ............................................................. 3

1.1.4. Hi chng mn kinh ..................................................................................... 4

1.2. Cc nghin cu v thnh phn ha hc v tc dng dc l ca cy

Mn kinh ( Vitex trifolia L.) ................................................................................ 5

1.2.1. Nghin cu v thnh phn ha hc .............................................................. 5

1.2.2. Nghin cu v tc dng dc l ................................................................ 10

1.3. Cng dng theo y hc c truyn ................................................................ 13

Chng 2: i tng v phng php nghin cu ........................................ 16

2.1. i tng ...................................................................................................... 16

2.2.1. i tng nghin cu ................................................................................ 16

2.1.2. Nguyn vt liu v trang thit b nghin cu ............................................. 16

2.2. Phng php nghin cu ............................................................................ 17

2.2.1.Vi hc .......................................................................................................... 17

2.2.2. Nghin cu thnh phn ha hc ................................................................. 17

2.2.3. Nghin cu tc dng dc l ..................................................................... 18

2.2.4. X l s liu ............................................................................................... 20

Chng 3: Kt qu v bn lun ........................................................................ 21

3.1. c im vi hc ca mn kinh t .............................................................. 21

3.1.1. c im vi phu ...................................................................................... 21

3.1.2. c im bt dc liu ............................................................................. 21

3.2. Thnh phn ha hc ca mn kinh t ...................................................... 23

3.2.1. Tinh du ..................................................................................................... 23

3.2.2. nh tnh cc nhm cht hu c khc trong mn kinh t .......................... 23

3.2.3. nh tnh bng phng php sc k lp mng .......................................... 29

3.3. Tc dng ca cao chit cn 90 mn kinh t i vi hnh vi lo lng v

ci thin tr nh/nhn thc ................................................................................ 31

3.3.1. Tc dng ca cao chit cn mn kinh t i vi hnh vi lo lng .............. 31

3.3.2. Tc dng ci thin tr nh / nhn thc ca cao chit cn mn kinh t ...... 32

3.3.3. Trng lng t cung vi trng ................................................................ 34

3.4. Bn lun ....................................................................................................... 35

KT LUN V XUT .......................................................................... .....37

LI CM N

c c thnh cng ca kha lun ny, li u tin em xin c by

t lng bit n su sc n TS. Phm Nguyt Hng Vin Dc liu, ngi

thy, ngi ch lun bn cnh ch bo, hng dn v gip em tn tnh

trong thi gian thc hin ti.

Em cng xin chn thnh cm n ThS. Phm Thi H Vn B mn

Dc hc c truyn, trng i hc Dc H Ni ngi thy lun ht lng

gip , to iu kin v ng gp nhng kin qu bu em hon thnh

tt kha lun ny.

Em xin chn thnh cm n cc thy c, anh ch B mn Dc hc c

truyn trng i hc Dc H Ni Khoa v Khoa Dc l Sinh ha

Vin Dc liu ng vin v gip em c v vt cht ln tinh thn em

c th hon thnh kha lun tt nghip.

Em xin chn thnh cm n Ban gim hiu, ng y nh trng cng

ton th cc thy c gio Trng i hc Dc H Ni trang b y

kin thc v ng vin, gip em trong sut qu trnh hc tp v nghin cu

ti trng.

Cui cng, em xin by t lng bit n su sc ti gia nh v bn b

lun l ch da tinh thn, l nhng ngun ng vin to ln i vi em

trong cuc sng cng nh trong hc tp.

H Ni, thng 5 nm 2014

Sinh vin

Trng Minh Phng

DANH MC CC CH VIT TT

DVN: Dc in Vit Nam

Dl: Dc liu

EtOH: Ethanol

EtOAc: Ethyl acetat

IC50: Nng c ch mc 50%

LC50: Nng gy cht mc 50%

MIC: Nng c ch ti thiu

Sham: Nhm chng sinh l

SKLM: Sc k lp mng

TT: Thuc th

UV: Ultra Violet spectroscopy ( Ph hng ngoi )

OVX: M hnh chut nht ci phu thut ct b bung trng 2 bn

VT: Cao chit cn ca qu Vitex trifolia ( mn kinh t )

XLC: Xanh l cy

XD: Xanh dng

X: Xanh en

DANH MC CC HNH

Hnh 1.1. Cy mn kinh Vitex trifolia L.

Hnh 1.2. Qu kh ca cy Mn kinh Vitex trifolia L. ( Mn kinh t )

Hnh 3.1. c im vi phu qu

Hnh 3.2. c im bt mn kinh t

Hnh 3.3. Sc k ca tinh du mn kinh t quan st di nh sng t ngoi

bc sng = 254nm (a), = 366nm (b) v sau khi phun TT vanillin (c).

Hnh 3.4. Sc k ca flavonoid trong mn kinh t quan st di nh sng

t ngoi bc sng = 254nm (a), = 366nm (b).

Hnh 3.5. Biu ct biu din t l thi gian chut cung phn t ch (%)

DANH MC CC BNG

Bng 1.1. Mt s thnh phn ha hc ca cy Mn kinh

Bng 3.1. Kt qu nh tnh cc nhm cht trong mn kinh t bng phng

php ha hc.

Bng 3.2. Kt qu nh tnh bng SKLM ca tinh du mn kinh t.

Bng 3.3. Kt qu nh tnh bng SKLM ca flavonoid t mn kinh t.

Bng 3.4. Tc dng ca cao chit cn mn kinh t trn th nghim nh gi

hot ng t nhin ca chut

Bng 3.5. Thi gian tim n tm thy platform

Bng 3.6. Tc dng ca cao chit cn mn kinh t trn trng lng t cung

vi trng ca chut OVX

1

T VN

Mn kinh t l v thuc thuc danh mc v thuc thit yu ca thuc y

hc c truyn. Theo kinh nghim dn gian, mn kinh t c tc dng pht tn

phong nhit, c s dng trong iu tr cm cm, au u, au mt mt

s nc Chu nh Thi Lan, n , Indonexia, Trung Quc, mn kinh t

cng c s dng ph bin trong iu tr mt s bnh nh au u, chng

mt, ti tm mt mi, hen suyn

S pht trin ca khoa hc k thut to iu kin thun li cho s

pht trin ca y hc hin i. Nhng kinh nghim s dng v thuc, bi thuc

ca Mn kinh t trong dn gian c lm sng t bng y hc hin i qua cc

cng trnh nghin cu v thnh phn ha hc, tc dng dc l ca chng.

Mn kinh t l v thuc c s dng kh ph bin theo kinh nghim dn

gian nhiu nc, c bit l cc nc Chu . Do cng c nhiu cng

trnh nghin cu v thnh phn ha hc, tc dng dc l ca Mn kinh t v

phn no chng minh c kinh nghim s dng trong iu tr bnh theo kinh

nghim dn gian. Theo y hc hin i, Mn kinh t c tc dng khng khun,

khng nm, tc dng trn mt s dng t bo ung th, lm lnh vt thng, c

hot tnh chng vim, gim au, khng Histamin

Tuy nhin cha thy c nghin cu no chng minh tc dng ph n

giai on mn kinh. gp phn nng cao gi tr s dng ca Mn kinh t,

chng ti tin hnh thc hin ti Nghin cu thnh phn ha hc v

tc dng gim lo lng, tng tr nh ca Mn kinh t ( Vitex trifolia L. )

theo cc mc tiu sau:

1. Xc nh c cc nhm cht ha hc chnh ca Mn kinh t.

2. Th nghim tc dng gim lo lng , tng tr nh ca Mn kinh t trn m

hnh OVX.

2

Chng 1: Tng quan

1.1. Thng tin chung

1.1.1. V tr phn loi ca chi Vitex

Cy Mn kinh thuc chi Vitex, mt chi thuc h C roi nga

Verbenaceae. V tr ca chi Vitex trong h thng phn loi thc vt c tm

tt nh sau [5]:

Gii: Plante

Ngnh: Mangnoliophyta

Lp: Magnoliapsida

Phn lp: Lamiidae

B: Lamiales

H: Verbenaceae

Chi: Vitex L.

Mn kinh cn c tn gi l Quan m, n ba l, T bi bin, Vn kim

t, Mc nim (Ty), Indian wild pepper (Anh)

1.1.2. c im chung ca chi Vitex

Chi Vitex c khong 150 loi phn b trn khp cc vng nhit i v

nhit i ca chu , chu Phi v chu M; ch c mt s t loi vng n

i m chu u v chu . Malyasia c khong 30 loi, n c hn 30

loi. Vit Nam c 14 loi, trong c 6 7 loi c dng lm thuc [11].

Nhng loi thuc chi Vitex l cy g hay cy bi. Cnh non vung hay

trn, nhn hay c lng. L mc i, kp, chn vt, thng 1- 3 - 5 - 7 l cht,

c mp nguyn hay x rng ca. Cung l thng trn, c rnh pha trn

hay c cnh. Cm hoa nh cnh hay nch l pha nh cnh; gm cc xim 2

ng v 3 ng hp thnh hnh chy, hnh thp hay hnh ng. L bc nh tn ti

hay sm rng. i hnh chung, hnh chn hay hnh ng, c 5 thy ngn, ct.

3

Trng c ng ngn, thng hay hnh phu, 2 mi: mi trn 2 thy, mi di 3

thy vi thy gia ln hn. Nh 4, thng 2 di, 2 ngn tht vo hay th khi

ng trng; ch nh nhn hay c lng pha di, nh trn ng trng; bao phn

2 thng dng ra. Bu 2 4 , mi 1 2 non, vi nhy nh, nh x 2

thy. Qu hch gn hnh cu hay hnh trng, nang i tn ti, v qu trong

ha g cng, v qu gia nc, thng cha 1 2 ht [10].

Mt s loi thuc chi Vitex : Vitex trifolia L., Vitex rotundifolia L.,

Vitex negundo L., Vitex agnus catus L.,

1.1.3. c im ca loi Vitex trifolia L.

Hnh 1.1. Cy Mn kinh Vitex trifolia L.

1.1.3.1. c im thc vt

- Cy nh hay cy bi, mi thm, c th cao ti 3m. Cnh non c 4 cnh, c

lng mm, mu xm nht; cnh gi trn, nhn, mu nu.

- L kp mc i, 3 l cht (l ngn c hoa thng n), phin l cht hnh

trng, gc trn, u t hoc hi nhn, mp nguyn, mt trn nhn hoc en

li khi kh, mt di ph y lng trng, l cht gia ln hn l cht hai bn,

gn khng ni r; l v ra c mi thm; cung gy, hi trn, c lng, di 1-

3cm, l cht khng c cung. Hoa mu l nht di 13 14 mm.

- Cm hoa l mt chy tn cng, i khi c l gc, c lng dy; mang nhiu

xim mc i, mi xim c 2-3 hoa mu tm nht hoc lam nht; l bc nh,

4

hnh di; i hnh chung, c lng trng, 5 rng nh u; trng hnh tr c

lng mt ngoi tr phn gc, mi trn c 2 thy ngn, mi di 3 thy, thy

gia ln hn hai thy bn; nh 4, th ra ngoi [9], [11].

Hnh 1.2. Qu kh ca cy Mn kinh Vitex trifolia L. ( Mn kinh t )

Mn kinh t l qu ca cy Mn kinh Vitex trifolia L. Qu hch, hnh

cu, ng knh 4 6 mm, mt ngoi mu nu en hoc xm en, ph lng

nhung mu xm nht nh sng, c 4 rnh dc nng, u hi dt, nh hi

lm, y c i tn ti mu xm nht v cung qu ngn. L i bao bc 1/3

2/3 qu, c rng, trong c 2 rng x tng i su, c ph kn lng t

mt. Cht nh v cng, kh p v. Mt ct ngang qu c 4 , mi c 1

ht, Mi thm c bit, v nht, hi cay. Ma hoa qu t thng 4 11 [3],

[11].

1.1.3.2. Phn b

Mn kinh c ngun gc t Nam Phi, vng phn b t Madagasca n

Xrilanca, Afghanistan, n , Myanma, nam Trung Quc, Nht Bn, xung

n cc nc vng ng Nam n vng bc c v pha ng Caledona

[11].

Vit Nam: phn b ri rc khp cc tnh cng ni thp xung n

trung du, i khi gp c ng bng. cao phn b thng < 1000m. Mc

hoang ch yu vng bin n rng ngp mn t Thanh Ha qua Nng

tr vo n Tin Giang ( G Cng ) v Kin Giang ( H Tin ) [6].

1.1.4. Hi chng mn kinh

Hi chng mn kinh ( menopause syndrome ) l tp hp cc triu

chng xut hin trong giai on mn kinh ph n do chc nng ca bung

5

trng suy thoi v hm lng hormone sinh dc n ( estrogen ) trong c th

h thp gy nn. thi k mn kinh, ngi ph n phi i din vi cc nguy

c cao v mc cc bnh nh tm thn kinh, tim mch, long xng, tuyn ni

tit, h vn mch, hi chng bin dng. C rt nhiu nghin cu ch ra v

hi chng tm thn kinh ca ph n giai on ny nh mt ng, lo lng, d

ni cu, d tht vng, trm cm, mt mi, gim tr nh

Nguyn nhn ca cc vn ny l do ni tit t n estrogen thay i

su sc [14]. Trc tui mn kinh, bung trng hot ng theo chu k hng

thng v tit ch ra cc nang non, ng thi phng thch cc hormone nh

estrogen v progesterone i vo mu lm c s cho th thai. tui mn kinh

cc nang trng nguyn y cn kit, bung trng ngng hot ng hn dn ti

mt hon ton ni tit t n l estrogen s dn ti gim v mt cn bng trong

mi tng quan gia chc nng tuyn yn, bung trng, tuyn gip, tuyn cn

gip, tuyn thng thngy nn cc triu chng ca hi chng mn kinh.

Khi mn kinh bung trng khng tit ra estrogen na, nhng tuyn thng

thn tit ra androstenedion, m m v mt s t chc khc ca c th chuyn

ha thnh estrogen duy tr nhng chc nng ngi ph n. Mt s

nghin cu ch ra mi lin quan gia estrogen, tr nh v h cholinergic.

Liu php iu tr hormone thay th i vi ph n mn kinh c tc dng lm

chm v gim nguy c mc cc triu chng trn v bnh Alzheiner [20], [30].

1.2. Cc nghin cu v thnh phn ha hc v tc dng dc l ca cy

Mn kinh ( Vitex trifolia L. )

1.2.1. Nghin cu v thnh phn ha hc

1.2.1.1. Trn th gii:

Cy mn kinh Vitex trifolia L. l cy thuc c s dng ph bin trong

nn y hc c truyn ca cc nc chu . Trung Quc, Nht Bn l hai nc

c nhng nghin cu u tin v thnh phn ha hc ca l v qu Vitex

6

trifolia L. Vitex trifolia L. l mt dc liu tim nng v vn ang tip tc

c nghin cu.

Bng phng php sc k, phn tch d liu quang ph, phn tch tinh th

bng tia X v phn ng ha hc, xc nh c trong l v qu Vitex

trifolia L. c cc thnh phn nh: tinh du ( cc terpenoid: pinen, linalool,

terpinyl acetat ), flavonoids, alkaloids, lignans,

Thnh phn ha hc ca l Vitex trifolia L.: ch yu l tinh du v

phn lp c 40 cht ca tinh du bao gm: pinene, linalol, terpinyl

acetate, caryophyllene oxide, -caryophyllene[43]. Trong mt nghin cu

gn y nht ca cc nh khoa hc n , tch v xc nh c mt

halimane diterpenoid : 13 hydroxyl 5(10),14 halimadien 6 one v hai

labdane diterpenoids: 6,7 diacetoxy 13 hydroxyl 8(9),14 labdadien

v 9 hydroxyl 13(14) labden 15,16 olide trong phn on n hexan

t dch chit u ca l Vitex trifolia trong methanol bng phng php TLC

vi pha ng l h dung mi chloroform/aceton (98:2 v/v) [41].

Thnh phn ha hc ca qu Vitex trifolia L. ( Mn kinh t ): thnh

phn chnh l tinh du; ngoi ra cn c flavonoid, lignan, lactone, acid hu

c, alkaloid, vitamin A

Tinh du: qua cc nghin cu phn lp c mt s

monoterpenenoid, diterpenenoids v triterpenenoids:

Monoterpenoid: camphen, pinen,

Diterpenoids: phn lp c rotundifuran, dihydrosolidagenone,

abietatriene 3 ol, Vitetrifolin A, B, C, D, E, F, G t dch chit acetone ca

qu Vitex trifolia L. [31] v vitetrifolin H, I [44].

Triterpenoids : ursolic acid; 2 , 3 dihydroxyurs 12 en 28 oic

acid; betulinic acid; taraxerol; 2 , 3 , 19 trihydroxyurs 12 en 28

oic acid[13].

7

Mt s thnh phn khc:

Theo Wehmer (1931, Die Pflanzenstoffe, Bd, 1023) th trong qu ca cy

Vitex trifolia L. c ankaloid v vitamin A.

Nhm phyosterol: sitosterol, sitosterol 3 O glucoside [43] v

stigmast 4 en 6 ol 3 one c phn lp t dch chit n buthanol

[16].

Acid hu c: rho-hydroxybenzoic acid [44].

Nhm lactone: viteosin A c phn lp t dch chit n-hexan ca qu

[12] v vitexilactone, previtexilactone c phn lp t dch chit aceton

[25].

Nhm flavonoid: casticin ( vitexcarpin), 3, 6, 7 trimethyl quercetagetin

[44], persicogenin, luteolin, penduletin, chrysosplenol D, artemetin [27].

Nhm lignan: vitrifol A ( dimer ca dihydrobenzofuran) v dihydro

dehydrodiconifenyl alcol phn lp c t dch chit n buthanol [16].

Cht to mu ca tinh du: xc nh c 2 norditne aldehydes t dch

chit acetone ca qu mn kinh thu hi ti thnh ph H Ch Minh thng

6/2000 [25].

1.2.1.2. Vit Nam

n nay c rt t nghin cu v thnh phn ha hc ca Mn kinh t.

- Theo cc ti liu trong nc, Mn kinh t c cc thnh phn chnh l tinh

du, flavonoid, alkaloid, vitamin A...[6], [11]

+ Tinh du gm c: L pinen v camphen (55%), terpinyl acetat (10%),

diterpen alcol (2%)Hm lng tinh du trong l mn kinh ti khong t

0,42 0,50 % v trong qu l 0,07 0,23 %.

+ Flavonoid gm c: aucubin, agnusid, casticin, orientin, luteolin 7

glucosid, iso orientin

+ Alcaloid: vitricin ( vitexin )

8

- Ht cha acid p hydrobenzoic, acid p anisic, vanillin

- L cha 0,28% tinh du ( tnh theo dc liu kh ), trong c terpinyl

acetat (10%), 1,8 cineol, pinen

Cng trnh nghin cu gn y nht v thnh phn ha hc ca tinh du

mn kinh c thc hin bi Nguyn Vn Bi ( i hc S phm, i hc

Hu ) c cng b trn Tp ch khoa hc i hc Hu, 22/6/2004. Bng

phng php sc k kh khi ph lin hp ( GC/MS ) xc nh c thnh

phn ha hc chnh trong tinh du l mn kinh l: 6-(1,2-dimetyl1

propenyl) 4,5 diazaspiro [2.4] hept 4 en (20,92%); 3 thujen

(15,85%); 1, 4 H, 10 H guaia 5,11 dien (13,40%); 3,7,11,15

tetrametyl hexadeca 1,3,6,10,14 pentane (6,98) v thnh phn ho hc

chnh ca tinh du qu mn kinh l: cis ocimen (7,29%); eucalyptol

(9,45%); axetat p menth 1 en 8 yl (5,56%); isocaryophyllen (9,04%)

[4].

Bng 1.1: Mt s thnh phn ca cy Vitex trifolia L.

STT Tn hp cht B phn Cng thc

1 pinen L, qu

2 Terpinyl acetat L, qu

3 Rotudinfuran Qu

9

4 Vitetrifolin A Qu

5 Dihydrodehydro

diconifenyl alcol Qu

6 Casticin

(Vitexcarpin) Qu

7 Luteolin Qu

8 3,6,7 trimethyl

quercetagetin Qu

10

9 Orientin Qu

10 p hydroxybenzoic

acid Ht

2.2. Nghin cu v tc dng dc l

Trn th gii c rt nhiu nghin cu v tc dng dc l ca cy

Vitex trifolia L. c cng b trn cc tp ch, bo co khoa hc vi cc tc

dng nh: h st nh, khng khun, khng virus, khng histamine, dit nm,

cn trng, tc ng trn dng t bo ung th

o Tc dng khng khun, khng nm:

- Phn on n hexan ca dch chit methanol t l Vitex trifolia L. thu c

2 terpenoids: 9 - hydroxyl - 13(14) labdien - 15,16 - olide v isoambreinolide

c tc dng vi vi khun lao Mycobacterium tuberculosis H37Rv trong th

nghim BACTEC 460 nng c ch ti thiu ( MIC ) ln lt l

100g/ml v 25 g/ml [40].

- Dch chit chloroform t l Vitex trifolia L. c tc dng khng khun vi

Pseudomonas aeruginosa, Klebsiella pneumonia v dch chit nc c tc

dng khng khun vi Staphylococus aureus nng 100 mg/ml [15].

- Dch chit cn v hexan ca l Vitex trifolia L. c tc dng c ch trn c vi

khun Gram m ( Klebsiellasp, Pseudomonas aeruginosa, Samonella typhi,

11

Escherichia coli, Shigella boydii) v Gram dng ( Bacillus cereus,

B.subtilis, B.megaterium, Sarcena lutea, Staphylococcus aureus ) [22].

- Dch chit hexan, dichloromethane, methanolca l c ch s sinh trng

ca mt s vi khun nh Staphylococus aureus, Streptococus faecalis,

Proteus mirabilis, E.coli, Shigella sonei ty theo nng tc dng [21].

- Dch chit hexan t l c ch hon ton s pht trin ca cc si nm

Furasium.sp trong 2 ngy u v gim dn sau 6 ngy [22].

o Tc dng trn t bo ung th:

- Dch chit hexan v dicloromethanic t thn v l cy Vitex trifolia L. gy

c vi mt s dng t bo ung th (SQC-1 UISO, HCT-15 COLADCAR,

KB, OVCAR-5) [21].

- 7 terpenoids: trifolin A G c chng minh c kh nng gy c trn mt

s t bo ung th ngi: A549, HCT 116, HL 60, 2R 75 30 vi gi tr

IC50 tng ng l 0,5; 0,6; 0,003; 0,002 (g/ml) bng phng php MTT

[45].

- Diterpenoids ( vitextrifolia H, I ) v monoterpenoid ( vitexoid ) c ch s

tng sinh t bo Hela vi gi tr IC50 trong khong 4 28 M. Vitextrifolin I

c ch chu k t bo pha G0/G1 v gy cht t bo t bo theo chng trnh

(apoptosis) [22].

- 6 flavonoids: aucubin, agnusid, orientin, luteolin -7- glucosid, iso -

orientin, casticin c tc dng c ch s gia tng ca t bo ung th ng vt

c v, c nh gi bng phng php SRB v tc ng trn chu k t bo

pha G2/M1, gy ra s tiu hy ca t bo, c th nghim trn dng t bo

tsFT210 [27].

- Vitexcarpin (casticin) c ch ng k s tng sinh ca cc t bo ung th

ngi, bao gm t bo A2780, HTC 15, HT 1080, K562 vi cc gi tr

12

IC50 tng ng l 19,12,4 M (48h), 0,660,1 M (48h), 0,440,006 M

(48h), 0,280,14M (24h) [41].

- Dch chit cn 95% ca mn kinh t thu c 5 labdane diterpenoids:

vitexilactone,(5S,6R,8R,9R,10S)-6-acetoxy-9-hydroxyl-13(14)-labden-16,15-

olide, rotundifuran, vitetrifolin D, vitetrifolin E c tc dng chng tng sinh

trn cc t bo ung th thng qua gy cm ng apoptosis trn t bo tsFT210

v K562 nng cao v c ch s tin trin ca chu k t bo pha G0/G1

v pha G2/M nng thp [26].

o Tc dng khc:

- Tc dng h st: phn on hexan t dch chit chloroform ca qu Vitex

trifolia L. c tc dng h st nh [18].

- c ch gii phng histamine: dch chit cn v hexan t l ca Vitex trifolia

L. c ch s gii phng histamine IgE t t bo RBL 2H3 [18].T dch

chit n hexan thu c hai thnh phn chnh l viteosin A v vitexcarpin c

tc dng lm n nh chc nng mng t bo mast, do c th ngn

histamine c gii phng t t bo mast [12].

- Hot tnh chng vim:

+ Dch chit nc t l c ch NF kB chuyn v thng qua biu hin ca NF

kB p50 ca tiu n v mRNA, nhng khng c tc dng trn p65. Do

gim nng cc cht trung gian ca phn ng vim: chemokine CCL 3,

CXCL 10, COX 2 [31].

+ Dch chit nc ca l c ch hot ng trn interleukin (IL) 1,6 v tng

hp iNOS mRNA v tc dng nh trn yu t hoi t khi u (TNF) [32].

- Lm lnh vt thng: dch chit cn ca l lm gim din tch vt thng

ng k: gim khong 72,35% sau 8 ngy iu tr [29].

- Trn virus: dch chit nc ca Vitex trifolia L. ( phn trn mt t ) cho

thy tc dng c ch virus HIV 1 RT l 98,06% nng 200g/ml [42].

13

- Tc ng trn u trng:

+ Methyl p hydroxybenzoate thu c t dch chit methanol ca l Vitex

trifolia L. gy cht 100% u trng ca Culex quinquefasciutus v Aedes

aegypti nng 20ppm vi gi tr LC50 tng ng l 5,77 v 4,74 ppm [24].

+ nh hng ca tinh du trn giai on u trng ca loi Spilosoma oblique

dao ng t 25 75% vi lng tinh du t 0,5 2,5l v c ch kh nng

sinh sn ca trng t 69,67 91,01 vi lng tinh du t 2,0 0,5l [37].

- Acid p hydroxybenzoic nng 10-3 c tc dng c ch men tyrosinase

t 71%, cn acid p amisic nng 10-3 t 34% [11].

- Thnh phn thu c t ct ko l mn kinh: bng ng tim tnh mch

vi liu lng 10mg/kg c tc dng i vi nhng ri lon tun hon kt

mc mt v mng treo rut do dextran gy nn nh nht ca mu tng cao,

ngng tp tiu cu, tc lu truyn mu gim. iu cho thy l mn kinh

c tc dng tng cng vi tun hon ngoi bin v ni tng [11].

1.3. Cng dng theo y hc c truyn

Theo ti liu c, mn kinh t c v cay, tnh hi hn, quy 3 kinh : can,

ph, bang quang; c tc dng pht tn phong nhit. Dng cha au u,

u nhc, mt hoa, mt au [6].

Hin nay: Mn kinh t c s dng trong iu tr mt s bnh nh:

cm mo phong nhit, nhc u, chng mt, nhc bn thi dng, gim au;

au mt, mt, nhiu nc mt, hoa mt, mt m nhn khng r; li rng

sng au; phong thp, gn ct au, t but; tiu ha khng bnh thng, vim

rut, a chy. L gi p ngoi cha n ng tn thng. Ht, l lm gi tr

au u [6].

Liu dng: ht 8 12 g / ngy, sc nc ung hoc 2 3 g / ngy, thuc bt.

King k: Nhc u, au mt do huyt h; ngi c v h.

Tng k: u, Thch cao.

14

Cch bo ch:

o Sy b tp cht, dng sng ( thng dng ).

o Tm ru sao qua dng trong trng hp phong thp c co git.

Mt s bi thuc: [9], [11]

Cha cm st, nhc u, mt sung : Mn kinh t 15g, Cc hoa 12g,

Chi t 12g, Bc h 12g, Kinh gii 10g, Xuyn khung 4g. Sc, xng cho ra

m hi, ung thuc khi cn nng .

Cha au u do huyt p cao ( Mn kinh t thang ): Mn kinh t 12g,

Cc hoa 12g, Bc h 8g ( cho sau ), Bch ch 8g, Cu ng 12 18 g.

Cha thin u thng: Mn kinh t 12g, Cc hoa 8g, Xuyn khung 4g, T

tn 3g, Bch ch 3g, Cam tho 4g. 600ml nc sc c cn 1/3 chia 3

ln, ung trong ngy.

Cha au mt sng , c mng che, chy d, qung mt: Mn kinh t,

ht Mung sao, ht Mo g trng, ht M , ht ch mu. Cc v bng nhau,

tn bt lm vin, ung vi nc ch hoc mi v 12g sc ung.

Tiu tin khng thng: Mn kinh t 10g, nghin bt chia 3 ln, ung vi

nc m trong ngy. Lm nhiu ln cho ti khi cc triu chng thuyn

gim.

Sng v: khi mi b, dng mn kinh t sao dn, tn nh. Mi ln ha 4g

vi ru. Gn ly ru ung cn b p ln v ( Bn tho cng mc ).

Lm cho tc en v di: Mn kinh t v m gu, 2 v bng nhau, trn vi

dm thanh bi vo tc.

Kinh nghim s dng trong dn gian mt s nc:

+ n : L p ngoi tr au thp khp, bong gn; l nn lm gi dng tr

vim chy v au u; l nghin bt dng tr st gin cch. Hoa dng km

vi mt ong tr st km theo nn v kht nhiu. Qu c dng tr b

15

kinh. L non mn kinh c tc dng khng khun v dit cn trng. Dch

hm t l c tc dng h st [6].

+ Indonexia: Cha bnh ph thng, au bng, l, a chy, au d dy v

rut, l lch sng to, bnh ngoi da, au ngc, ho, nhc u v st [6].

+ Thi Lan: L c dng tr bnh ngoi da v gh nga. R tr bnh au

gan v tr st. Qu tr hen suyn v tr [6].

+ Trung Quc: S tn phong nhit. Tr au u, au nu rng, au mt

v nhiu nc mt, chng mt, mt mi ti tm. Liu dng: 3 10 g [9].

Bo ch theo Trung y: dng mn kinh t th:

- B tai, tm ru mt lc, chn 3 gi, phi kh dng ( Li Cng ).

- B tai, gi nt dng ( L Thi Trn ).

Ti mt bnh vin thuc tnh Phc Kin, ngi ta th nghim s dng

mn kinh t chng th thai theo cch nh sau: Mn kinh t 30g, g sng

1 con. em nu chn, n ci v ung nc; sau khi sch kinh hoc sau khi

.

Theo mt s ti liu, Trung Quc s dng l v qu Vitex trifolia L.

trong iu tr ung th.

16

Chng 2: i tng v phng php nghin cu

2.1. i tng

2.2.1. i tng nghin cu

Mu nghin cu: qu ca cy mn kinh Vitex trifolia L. ( Mn kinh t )

c thu hi vng ni Ngh An, vo thng 4 nm 2013. Loi b tp, phi

kh.

Mu cy ti c Khoa Ha Thc vt Vin dc liu gim nh tn

khoa hc l Vitex trifolia L., h C roi nga Verbenaceae v c lu tr ti

khoa Ha Thc vt Vin dc liu, Vit Nam.

2.1.2. Nguyn vt liu v trang thit b nghin cu

ng vt nghin cu:

Chut nht trng ci khe mnh 8 10 tun tui, trng lng 18 20 g,

cung cp bi Hc vin Qun y.

Dung mi, ha cht:

Dung mi: EtOH 90, EtOAc, ether du ha, acid formic, toluene, nc.

Thuc th: dd FeCl3 5%, dd NaOH 10%, dd gelatin 1%, anhydrid acid,

acid picric 1%, bt Na2CO3, bt Mg, H2SO4 c, HCl c, NH3 c, TT

Bouchardat, TT Mayer, TT Dragendroff, TT Fehling A & B. TT KOH/EtOH

90, TT vanillin 1% trong dd acid sulfuric.

Ha cht khc: cn st trng, dung dch betadine 10%.

Thuc: Progynova ( estradiol valerate ), Diethyl ether.

Bn mng: bn mng trng sn silicagel 60 F254 c dy 0,2 mm ca

hng Merkc.

Dng c, trang thit b

B dng c ct tinh du ci tin.

B dng c ct thu hi dung mi.

17

Cn k thut PRESICA.

n soi UV Camag = 254 nm, = 366 nm.

Knh hin vi Labonad.

T sy SHELLAB.

Cc dng c: bnh cu, pipet, l thy tinh, ng nghim, kim tim, u cn,

gang tay, m,

B thy tinh hnh trn ( m b ), Platform.

2.2. Phng php nghin cu

2.2.1.Vi hc

Nghin cu c im vi hc theo phng php s dng knh hin vi.

2.2.1.1. c im vi phu

Tin hnh lm tiu bn vi phu theo cc bc sau:

Chn qu kh thch hp.

Ct ngang qu bng dao lam sao cho lt ct mng v y 3 phn ( v

qu ngoi, v qu trong v v qu gia). Lm tiu bn bng nc ct.

Bc lp o bn ngoi ht. Lm tiu bn bng nc ct.

Quan st di knh hin vi, m t c im gii phu, chp nh bng my

nh qua knh hin vi.

2.2.1.2. c im bt dc liu

Quan st trc tip, ngi, nm xc nh mu, mi, v.

Ln tiu bn bt dc liu bng nc ct. Quan st, m t v chp nh

nhng c im in hnh ca bt qua knh hin vi.

2.2.2. Nghin cu thnh phn ha hc

Ct tinh du bng phng php ct ko hi nc.

nh tnh cc nhm cht hu c bng:

18

+ Phn ng ha hc: flavonoid, coumarin, alkanoid, anthranoid, tannin,

glycoside tim, saponin, acid hu c, acid amin, ng kh.

+ Sc k lp mng: nh tnh tinh du v flavonoid.

2.2.3. Nghin cu tc dng dc l

2.2.3.1. B tr th nghim

ng vt c chia thnh 5 nhm:

Nhm chng bnh l ( nhm m hnh OVX ): chut c phu thut ct

bung trng hai bn v c ung nc trong sut qu trnh nghin cu.

Nhm chng sinh l ( Sham ): chut c phu thut nh nhm gy m

hnh nhng khng ct bung trng hai bn v c ung nc trong sut qu

trnh nghin cu.

Nhm ung cao chit cn 90 mn kinh t liu tng ng 1,6 g dl/kg.

( OVX + VT 1,6 g/kg )

Nhm ung cao chit cn 90 mn kinh t liu tng ng 3,2 g dl/kg.

( OVX + VT 3,2 g./kg )

Nhm ung thuc chun Progynova liu 0,0005 g/kg.

( OVX + Progynova 0,0005g/kg ).

2.2.3.2. Tin hnh

Phu thut ct bung trng hai bn ( gy m hnh OVX ):

Bc 1: Gy m chut bng diethyl ether.

Bc 2: Chut nm sp, bc l v st trng vng m.

Bc 3: M bc l v ct c 2 bn bung trng. ng m gn dc theo

hai bn ct sng, di 1cm, cch ct sng 0,5cm, cch gc ui 3cm. Qua da,

lpdi da, lp c, gt b lp t chc m ta s thy bung trng chut c

mu trng hng, kch thc 2 3mm, hnh a cung.

Bc 4: Khu t chc lin kt, cn c thnh bng, lng theo lp; khu da

v st trng ti ch bng dung dch betadine 10%.

19

Bc 5: Chut sau phu thut c nh du ri a vo chung m v

theo di sau m n khi ng vt tnh to tr li. Sau m 7 ngy chut c th

a tr li nui chung vi cc chut khc trong nhm v c iu tr nh

trnh by trn.

2.2.3.3. Tiu ch nh gi

c tin hnh sau 10 12 tun phu thut v iu tr thuc, bao gm:

nh gi tc dng gim lo lng bng phng php theo di hot ng t

nhin ca chut bng my m hng ngoi: Hai tun sau khi phu thut,

chut c chia thnh 5 l nh trong phn b tr th nghim. Tip theo, chut

c iu tr bng mu nghin cu v thuc chun Progynova trong 1 tun.

Ngy th 8 sau khi chut ung mu nghin cu/ progynova/ nc s c

nh gi kh nng hot ng t nhin bng my m hng ngoi. So snh kh

nng hot ng ca chut gia cc nhm i chng v nhm nghin cu.

nh gi hnh vi ci thin tr nh bng th nghim m l nc Morris:

Mt tun sau khi nh gi kh nng vn ng, chut tip tc c nh gi

hnh vi ci thin tr nh/nhn thc bng th nghim m l nc Morris.

Th nghim m l nc Morris c thc hin theo phng php ca

Morris c sa i (Morris 1984). Ni dung ny c thc hin ti m l

nc Morris. M l l mt b hnh trn, c ng knh 1,1 m, chiu cao 30

cm. Lng nc trong m b duy tr mc 15 cm, nhit nc khong 20oC

trong sut qu trnh th nghim. Platform c t trong phng c t sn cc

hnh nh nh hng khng gian cho chut trong qu trnh hc tp. V tr

ca cc hnh nh ny khng c thay i trong sut qu trnh chut hc tp.

Mt platform c ng knh 7,5 cm, lm bng nha trong sut, c t ti

mt gc phn t ca m l. Ton b thi gian chut cn phi hc l 8 ngy,

bao gm:

Bi tp c platform:

20

Bi tp ny c tin hnh trong 7 ngy, chut s c c luyn tp nh

khng gian (training trial) khi platform c t di mt nc 1cm. Mi

ngy chut phi tp 3 ln, mi ln chut c t vo mt trong 3 gc phn t

cn li ca m l. Nu trong 60 giy chut khng tm c platform, chng s

c hng dn tm platform. Khi n c platform, chut s c 20

giy. Khong cch gia cc ln tp l 1 pht, trong thi gian ny chut nhanh

chng c lau kh v si m. Ton b qu trnh chut tp bi c ghi li

bng camera, s liu s c tnh ton bit c thi gian tim n chut c

th tm thy platform. S liu mi ngy s c tnh bng gi tr trung bnh ca 3

ln luyn tp i vi mi chut.

Bi tp khng c platform:

n ngy th 8 sau bi tp nhn thy platform, chut s c tp th li

tr nh (probe trial). Trong bi tp ny, platform c b ra ngoi, chut c

bi trong m l 1 ln duy nht trong 60 giy. Chut s nh li v tr ca

platform v c xu hng bi lu hn ti gc mt phn t ca m l c

platform c t t nhng ngy tp trc. Camera s ghi li v phn tch thi

gian chut bi ti mi gc phn t ca m l.

Cn trng lng t cung vi trng:

Chut sau khi nh gi hnh vi s b git, bc tch t cung vi trng

v cn trng lng.

2.2.4. X l s liu

Kt qu thc nghin c biu din di dng: gi tr trung bnh cng/tr

lch chun SD: M SD (M: gi tr trung bnh tng l, SD: lch chun).

So snh gi tr trung bnh tng l gia cc mu bng phng php One

way ANOVA, s dng Dunnet test so snh v gia cc l th vi l

chng.

Kt qu c xem l c ngha thng k khi p < 0,05.

21

Chng 3: Kt qu v bn lun

3.1. c im vi hc ca mn kinh t

3.1.1. c im vi phu

V qu ngoi: gm 2 lp biu b v h b. Biu b c 1 lp cutin kh dy,

ri rc c lng tit hnh cu. H b t bo di, dt, thnh cng tng i kh dy.

V qu gia: pha ngoi t bo khng ng u, hnh nhiu cnh, bu

dc hoc trn, thnh mng; pha trong t bo di xp dc, thnh dy ln hn.

V qu trong: cu to bi t bo m cng hnh ch nht hoc bu dc,

thnh rt dy, cng vo pha trong thnh t bo cng dy.

V ht cu to bi 1 2 lp t bo hnh mng.

(1) (2) (3) (4) (5) (6)

Hnh 3.1. c im vi phu qu

1. Lng tit, 2. Biu b, 3. H b, 4. V qu gia, 5. V qu trong, 6. V ht

3.1.2. c im bt dc liu

o Bt mu nu xm, c mi c bit, v cay.

o T bo biu b ca l i hnh hi trn, thnh t bo thng ln sng.

o Lng che ch c 2 - 3 t bo, t bo nh ln hn, c hnh bu.

22

o T bo v qu ngoi hnh nhiu cnh, c ng vn k ca cutin v vt tch

lng rng, c lng tit v lng che ch:

+ Lng tit c 2 loi, loi lng n bo u v 1 - 2 t bo chn v loi

lng c 2 - 6 t bo u v 1 t bo chn.

+ Lng che ch c 2 - 4 t bo, di 14 - 68 m, thng cong thnh nm li.

o T bo v qu gia hnh hi trn hay bu dc, thnh hi ha g, c l r.

o T bo tit thng b v, c cha cc cht tit, t bo k bn cha git du

mu vng nht.

o T bo m cng ca v qu trong hnh bu dc hay hnh vung, ng knh

10 - 35 m.

o T bo v ht hnh trn hoc hi trn, ng knh 42 - 35 m, thnh c vn

li,

ha g.

Hnh 3.2. c im bt mn kinh t

1. T bo v qu ngoi: 2. Lng tit, 3. Lng che ch; 4. Lng che ch; 5. T

bo v qu gia; 6. T bo m cng; 7. T bo v ht; 8. T bo biu b

2

5

1

1

2

5

3

4

6 7 8

23

3.2. Thnh phn ha hc ca mn kinh t

3.2.1. Tinh du

o Mu nghin cu: Qu kh ca cy Mn kinh xay nh bng thuyn tn.

o Tin hnh:

Dng c: B dng c ct tinh du ci tin ( da trn c s dng c theo

quy nh ca DVN I, 1971) ca TS. Nguyn Th Tm b mn Dc liu,

trng i hc Dc H Ni; s dng bnh hng cho tinh du nh hn nc.

Tin hnh: Cn 20 g mn kinh t c tn nh bng thuyn tn. Cho

dc liu vo bnh cu dung tch 500 ml c c nt mi. Thm 200 ml nc

ct. Lp b dng c. un si trong 5h bng bp in. Tt bp, ngui, ly

ng hng tinh du, gn phn tinh du pha trn nc sao cho khng b ln

nc.

o Kt qu: Tinh du thu c t qu mn kinh c t trng nh hn nc,

mu vng sng, c mi thm nng, hc; v ng, the.

3.2.2. nh tnh cc nhm cht hu c khc trong mn kinh t

Cho 5g dc liu vo bnh nn, thm 50ml EtOH 90, un hi lu cch

thy n si trong 10 pht, lc nng. Dch lc thu c em lm phn ng

nh tnh flavonoid, coumarin.

nh tnh flavonoid:

o Phn ng Cyanidin:

Cho vo ng nghim 2ml dch chit cn, thm mt t bt magie v nh

4 5 git dung dch HCl c. un cch thy trong 5 pht.

Kt qu: dch chit chuyn sang mu cam ( phn ng dng tnh ).

o Phn ng vi FeCl3 5%:

Cho vo ng nghim 2ml dch chit cn, sau thm 2 3 git dung

dch FeCl3 5%.

Kt qu: dch chit chuyn sang mu xanh en ( phn ng dng tnh ).

24

o Phn ng vi dung dch kim

Cho vo ng nghim 2ml dch chit cn, sau thm vi git dung dch

NaOH 10%. Xut hin kt ta vng. Tip tc thm 1ml nc ct, lc u ln.

Kt qu: ta tan ht v mu vng ca dung dch m ln (phn ng dng

tnh).

o Phn ng vi hi NH3

Nh 1 2 git dch chit cn ln hai ming giy lc, sy nh n kh.

Quan st di nh sng thng thy mu vng. em h mt ming giy lc

khc ln ming l ammoniac c m np.

Kt qu: mu vng ca dch chit trn ming giy lc em h ln ming l

ammoniac c m hn ( phn ng dng tnh ).

Kt lun s b: mn kinh t c flavonoid.

nh tnh coumarin:

o Phn ng m v ng vng lacton

Cho vo 2 ng nghim nh, mi ng 1ml dch lc

- ng nghim 1: thm 0,5ml dung dch NaOH 10%.

- ng nghim 2: nguyn.

un c 2 ng nghim trn ni cch thy n si.

Quan st thy:

- ng nghim 1: xut hin ta c vng.

- ng nghim 2: trong sut.

Thm vo c 2 ng, mi ng 1ml nc ct, lc u.

- ng 1: ta tan, dung dch trong sut, mu vng ca dung dch m ln.

- ng 2: khng xut hin ta c.

Thm vo ng 1 mt vi git dung dch HCl c.

Kt qu: khng thy xut hin ta c ( phn ng m tnh ).

o Phn ng hunh quang

25

Nh dch chit ethanol ln giy lc, sau nh chng 1 git dung dch

NaOH 10%. kh, che mt na mu vt cht th bng mt mnh kim loi

ri em soi di nh sang t ngoi = 366 nm trong mt pht, sau b

vt che ra. Quan st.

Kt qu: khng c s khc bit v cng mu gia hai na vt cht th

( phn ng m tnh ).

Kt lun s b: mn kinh t khng c coumarin.

nh tnh glycoside tim

Cn 5g dc liu cho vo bnh nn dung tch 250ml. Thm 100ml cn

25 ri ngm trong 24h. Gn dch chit vo cc c m dung tch 100ml. Tip

tc loi tp bng ch acetat 30%, lc loi tp. Dch lc c chuyn sang bnh

gn, chit ly glycoside tim bng cch lc vi chloroform ( hai ln, 8ml/ln ),

lc k tch lp. Gn lp chloroform vo mt cc c m sy kh. Chia

u dch vo 2 ng nghim nh c sy kh, bc hi dung mi trn ni

bp cch thy n kh. Cn thu c lm cc phn ng ha hc.

o Phn ng Libermann Boruchard

Cho vo ng nghim c cha cn 1m anhydride acetic, lc u cho tan

ht cn. t nghing ng nghim 45, cho t t theo thnh ng 1ml dung dch

acid H2SO4 c, trnh xo trn cht lng trong ng.

Kt qu: khng xut hin vng trn b mt tip xc gia hai lp cht

lng ( phn ng m tnh ).

o Phn ng Baljet

Pha thuc th Baljet: cho vo ng nghim dung dch acid picric 1% :

NaOH 10% ( 1: 9 ) lc u.

Cho vo ng nghim c cha cn 0,5ml dung dch Et2OH 90%, lc

u cho tan ht cn. Thm 0,5ml dung dch thuc th Baljet mi pha. So vi

ng chng ( khng c cn ).

26

Kt qu: dung dch c mu vng chanh, khng c mu cam ( phn ng m

tnh ).

Kt lun s b: mn kinh t khng c glycoside tim.

nh tnh alkaloid:

Cn 5g dc liu cho vo bnh nn dung tch 250ml. Thm 20ml dung

dch acid sulfuric 1N. un n si, ngui, lc. Kim ha dch lc bng

NH3 c (dng ch th vn nng ) n pH 9 10. Sau cho dch lc kim

ha vo bnh gn, lc vi chloroform 3 ln, gp dch chit chloroform li, loi

nc bng Na2SO4 sau lc vi dung dch acid sulfuric 1N 3 ln, mi ln

5ml. Ly dch chit acid cho vo 3 ng nghim, mi ng nghim 2ml. Nh

vo ng nghim 2 3 git ln lt cc thuc th sau:

ng 1: thuc th Mayer

Kt qu: xut hin kt ta trng ( phn ng dng tnh ).

ng 2: thuc th Dragendorff

Kt qu: xut hin kt ta vng ( phn ng dng tnh ).

ng 3: thuc th Bouchardat

Kt qu: xut hin kt ta vng nu ( phn ng dng tnh ).

Kt lun s b: mn kinh t c alkaloid.

nh tnh anthranoid: Phn ng Borntraeger

Cn khong 3g dc liu vo bnh nn dung tch 100ml, thm 20ml

dung dch acid sulfuric 1N, un si cch thy trong 15 pht. ngui, lc

qua giy lc gp np, ly dch lc cho vo bnh gn dung tch 100ml. Chit

bng 5ml chloroform, gn ly lp chloroform cho vo ng nghim, bc hi

dung mi cn khong 1ml, thm 1ml ammoniac 10% lc u, khng thy xut

hin mu hng lp trn, tip tc thm vo git NaOH 10%.

Kt qu: lp nc khng xut hin mu hng ( phn ng m tnh ).

Kt lun s b: mn kinh t khng c anthranoid.

27

Sau khi ct tinh du, lc dch chit nc qua giy lc gp np. Dch lc

lm cc phn ng sau:

nh tnh saponin: Phn ng to bt

Cho vo ng nghim to 10ml dch lc v lc mnh theo chiu dc ng

trong 5 pht, yn quan st.

Kt qu: Ct bt xut hin nhng khng bn, tan ngay trong khong 1 pht

( phn ng m tnh ).

Kt lun s b: mn kinh t khng c saponin.

nh tnh tannin:

ng 1: 2ml dch lc, thm 2 git FeCl3 5% (TT).

Kt qu: xut hin kt ta xanh en ( phn ng dng tnh ).

ng 2: 2ml dch lc, thm 2 git ch acetat 10% (TT).

Kt qu: xut hin kt ta bng trng ( phn ng dng tnh ).

ng 3: 2ml dch lc, thm vo git dung dich gekatin 1% (TT).

Kt qu: khng xut hin kt ta trng ( phn ng m tnh ).

Kt lun s b: mn kinh t khng c tannin.

nh tnh acid amin:

Cho vo ng nghim nh khong 2ml dch lc, thm vi git thuc th

ninhydrin 3%, un cch thy 3 pht.

Kt qu: khng thy xut hin mu xanh tm ( phn ng m tnh ).

Kt lun s b: mn kinh t khng c acid amin.

nh tnh acid hu c:

Cho vo ng nghim nh khong 2ml dch lc, thm mt t tinh th

Na2CO3.

Kt qu: xut hin bt kh ( phn ng dng tnh ).

Kt lun s b: mn kinh t c acid hu c.

nh tnh ng kh:

28

Thm 1ml dung dch thuc th Fehling A + 1ml dung dich thuc th

Fehling B vo ng nghim cha 2ml dch lc. un si cch thy 3 pht.

Kt qu: khng xut hin kt ta gch.

Kt lun s b: mn kinh t khng c ng kh

Bng 3.1. Kt qu nh tnh cc nhm cht trong mn kinh t bng phng

php ha hc

STT Nhm cht Phn ng nh tnh Kt qu Kt lun s b

1 Flavonoid

Cyanidin +

C FeCl3 5% +

dd kim +

NH3 c +

2 Coumarin M, ng vng lacton -

Khng c Hunh quang -

3 Alkaloid

Mayer +

C Dragendorff +

Bouchardat +

4 Anthranoid Borntraeger - Khng c

5 Glycoside tim Libermann Boruchard -

Khng c Baljet -

6 Saponin To bt - Khng c

7 Tannin

FeCl3 5% +

Khng c Ch acetat 10% +

dd gelatin 1% -

8 Acid hu c Na2CO3 + C

9 Acid amin TT Ninhydrin 3% - Khng c

10 ng kh Fehling - Khng c

29

Hnh 3.3. Sc k ca tinh du

mn kinh t quan st di nh

sng t ngoi bc sng =

254nm (a), = 366nm (b) v sau

khi phun TT vanillin (c).

Ch thch: (-) phn ng m tnh, (+) phn ng dng tnh.

Nhn xt: Ngoi tinh du, mn kinh t cn cha flavonoid, alkaloid, acid

hu c.

3.2.3. nh tnh bng phng php sc k lp mng

3.2.3.1. Tinh du

Chun b dch chm sc k: 1git tinh du c pha long trong 1ml ether

du ha ( 60 90 ).

Bn mng: Silicagel G hot ha 100C trong khong 1 gi.

Dung mi khai trin: ether du ha EtOAc ( 9/1 ) [8].

Quan st di nh sng t ngoi ti bc sng = 366 nm.

Hin mu: bng dung dch vanilin 1% trong acid sulfuric ( TT ), sy bn

mng 110C trong 10 pht.

Kt qu: Theo Bng 3.2

Khi cha phun thuc th:

- Quan st di nh sng t ngoi bc sng

=254nm : c 10 vt

( Rf = 1, 5, 7,21,23, 27, 33, 39, 55, 60 ).

- Quan st di nh sng t ngoi bc sng

=366nm : c 8 vt

( Rf = 1, 5, 7,21,23, 27, 33, 39 ).

Sau khi phun TT vanillin 1% quan st di nh

sng thng: c 10 vt

( Rf = 1, 5, 7,21,23, 27, 33, 39, 55, 60 ).

a b

c

30

Trong cc vt, nhn thy mt s vt ng ch :

- Vt 9 ( Rf100 = 55) v vt 10 ( Rf100 = 60) ch quan st c ti UV254nm

m khng quan st c ti UV 366nm, hin mu vng nu v nu sau khi phun

TT vanilin 1% ( quan st nh sng thng ).

- Vt 6 ( Rf100 = 27) hin mu r nht, c UV254nm v UV366nm, hin mu

xanh en sau khi phun TT vanillin 1% ( quan st nh sng thng ).

Bng 3.2. Kt qu nh tnh bng SKLM ca tinh du mn mn kinh t

Ghi ch: +: rt m; ++: m; +++: r; ++++: rt r.

3.2.3.2. Flavonoid

Chun b dch chm sc k: dch chit cn 90 ca mn kinh t.

Bn mng: Silicagel G hot ha 100C trong khong 1 gi.

Dung mi khai trin: Toluen : EtOAc : acid formic ( 5 / 4,5 /1).

Quan st di nh sng t ngoi ti bc sng = 254nm, = 366 nm

STT Rf 100 Mu sc

UV254nm UV366nm Hin mu

1 1 Xanh +++ XD +++ Nu

2 5 X ++++ XD +++ Nu

3 7 Xanh ++ XD +++ Nu

4 21 Xanh + XD +++ Xanh en

5 23 Xanh m

++++ XD ++ Nu

6 27 XD ++++ XD ++++ Xanh en

7 33 X +++ X + Vng nu

8 39 X ++ X ++ Tm

9 55 X +++ Vng nu

10 60 X ++++ Nu

31

Kt qu:

Bng 3.3. Kt qu nh tnh bng SKLM ca flavonoid

t mn kinh t

STT Rf100 Mu

UV254nm UV366nm

1 1 X +++ XD +++

2 30 en +

3 41 X ++ XLC ++++

4 48 X +++ XD ++++

Ghi ch: +: rt m; ++: m; +++: r; ++++: rt r.

- Quan st di nh sng t ngoi bc sng

=254nm: c 3 vt ( Rf = 1, 41, 48 ).

- Quan st di nh sng t ngoi bc sng

=366nm : c 4 vt ( Rf = 1, 30, 41, 48).

3.3. Tc dng ca cao chit cn 90 mn kinh t i vi hnh vi lo lng v

ci thin tr nh/nhn thc

3.3.1. Tc dng ca cao chit cn mn kinh t i vi hnh vi lo lng

nh gi tc dng gim lo lng bng phng php theo di kh nng

vn ng ca chut bng my m hng ngoi.

Kt qu bng 3.4 cho thy khng c s khc bit v kh nng vn

ng t ngha thng k gia nhm OVX so vi nhm Sham hoc OVX +

VT (1,6 g/kg ) hoc OVX + progynova. Tuy nhin, gim kh nng vn ng

nhm OVX + VT ( 3,2 g/kg ) so vi nhm OVX, t ngha thng k ( p <

0,05 ).

Hnh 3.4. Sc k ca

flavonoid t mn kinh t

quan st di nh sng t

ngoi bc sng =

254nm (a), =366nm (b)

a b

32

Bng 3.4. Tc dng ca cao chit cn mn kinh t trn th nghim nh gi

hot ng t nhin ca chut

*: p < 0,05 so vi nhm OVX

3.3.2. Tc dng ci thin tr nh/nhn thc ca cao chit cn mn kinh t

nh gi trn thi gian tim n tm thy platform:

Bng 3.5. Thi gian tim n tm thy platform

L th

nghim

Thi gian tim n tm thy platform (s)

Ngy 1 Ngy 2 Ngy 3 Ngy 4 Ngy 5 Ngy 6 Ngy 7

Sham 33,12,6 27,44,4 23,74,1 17,73,9* 15,02,5** 15,83,0** 14,32,5**

OVX 35,73,2 35,55,3 32,45,3 32,65,6 27,37,0 28,87,9 23,74,7*

OVX + VT

1,6 g/kg 31,84,8 24,32,9 33,44,7 28,31,6 25,82,4 20,79,0* 21,03,4

OVX + VT

3,2 g/kg 37,63,6 23,23,1 25,04,6 26,44,4 20,64,2 18,63,2 18,83,7*

OVX +

Progynova 30,93,6 26,93,2 28,23,0 25,44,1 23,64,9* 23,24,9 19,25,8*

*: p < 0,05; **: p < 0,01 so vi ngy 1 trong cng mt nhm

L th nghim N Liu (g/kg) Thi gian vn ng (s)

Sham 10 - 119,25 7,15

OVX 11 - 120,01 6,39

OVX + VT 11 1,6 111,48 8,14

OVX + VT 11 3,2 104,29 8,45*

OVX + Progynova 11 0,0005 117,71 11,61

0.0

10.0

20.0

30.0

40.0

50.0

1 2 3 4 5 6 7

Sham OVX OVX + Progynova OVX + VT 1,6 g/kg OVX + VT 3,2 g/kg

Th

i

gia

n t

im

n

(gi

y)

Ngy

33

bi tp khng c platform: chut cc nhm khc nhau u c kh

nng tm thy platform nm di mt nc 1 cm, vi thi gian tim n ngn

dn cc ngy luyn tp k tip. nhm Sham, gim thi gian tim n c

ngha thng k t ngy th 4 n ngy th 7 ( p < 0,01; p < 0,05 ). Thi gian

tim n ca nhm OVX cng gim qua cc bi tp, tuy nhin cn thi gian

di hn tm thy platform so vi nhm Sham (p < 0,05 ngy th 7). Thi

gian tim n ca nhm OVX + VT v OVX + Progynova cng c rt

ngn t ngy th 7 qua cc ngy tp (p < 0,05 ngy th 7). Nu so snh thi

gian tim n tm thy platform gia cc nhm ngy th 7, kt qu cho

thy nhm Sham, OVX + VT v OVX + Progynova c thi gian tim n

tm thy platform ngn hn nhm OVX.

T l thi gian chut cung phn t ch ( % )

Sau 7 ngy luyn tp, n ngy th 8, chut c nh gi kh nng

cng c li tr nh.

Hnh 3.5. Biu ct biu din t l thi gian chut cung phn t ch (%)

*p

34

bi tp ny, platform c ly ra khi b bi. Thi gian chut

cung phn t ch (cung phn t m trong nhng bi tp trc c platform) s

c ghi li nh gi kh nng cng c tr nh ca chut.

Kt qu hnh 3.5 cho thy: nhm chut OVX c t l thi gian

cung phn t ch t hn so vi nhm Sham (p < 0,05). Chut OVX c

ung Progynova hoc cao chit VT (1,6 g/kg v 3,2 g/kg) c t l thi gian

cung phn t ch nhiu hn so vi nhm chut OVX v gn tng ng

vi nhm Sham.

Nh vy, cao chit VT vi liu 1,6 g/kg v 3,2 g/kg c tc dng ci

thin tr nh/nhn thc trn chut OVX

3.3.3. Trng lng t cung vi trng

Kt thc cc bi tp nh gi hnh vi sau 2 tun iu tr, s bc tch t

cung vi trng ca chut v cn trng lng.

Bng 3.6. Tc dng ca cao chit cn mn kinh t trn trng lng t cung

vi trng ca chut OVX

**: p < 0,01; *: p < 0,05 so vi nhm OVX

Kt qu c trnh by bng 3.6 v cho thy: sau 2 tun c iu

tr bng mu nghin cu/thuc chun, trng lng t cung vi trng ca

chut u tng. Trong , nhm OVX + Pronovgya tng trng lng t cung

L th nghim N Liu

(g/kg)

Trng lng t cung

vi trng (mg %) % tng

Sham 13 - 139,26 15,49 -

OVX 11 - 78,26 8,94 -

OVX + VT 9 1,6 134,67 19,66* 72,08

OVX + VT 9 3,2 117,65 15,54* 50,33

OVX + Progynova 10 0,0005 366,69 50,76** 359,24

35

ln n 359,24% ( p < 0,01 ), nhm OVX + VT 1,6 g/ kg v 3,2 g/kg c

trng lng t cung vi trng tng 72,08 % v 50,33 % ( p < 0,05 ) so vi

nhm OVX.

Nh vy, nhm OVX + VT 1,6 g/kg v 3,2 g/kg c tc dng lm tng

trng lng t cung vi trng.

3.4. Bn lun

Trong giai on mn kinh ph n, chc nng ca bung trng suy

thoi v hm lng hormone sinh dc n ( estrogen ) trong c th h thp gy

nn cc triu chng nh bc ha, mt ng, lo lng, stress, d tht vng, trm

cm, ni cu, mt mi, gim tr nh.Theo kinh nghim dn gian, mn kinh

t c s dng iu tr cc bnh nh cm mo phong nhit, au u,

chng mt, nhc bn thi dng, ti tm mt mi, au mt, mt, nhiu

nc mt, hoa mt, mt m nhn khng rDo mn kinh t c th c tc

dng trong iu tr mt s triu chng ca hi chng mn kinh.

Trong chi Vitex c 2 loi l Vitex agnus catus L. v Vitex rotundifolia

L. c nghin, nh gi tc dng nh estradiol v giai on mn kinh.

T dch chit chloroform ca qu Vitex agnus catus L. phn lp c

casticin, vitexilactone, pinnatasterone and 17 OH progesterone v th tc

dng dc l trn m hnh OVX thu c kt qu: dch chit chloroform t

qu tng ng k trng lng t cung v tng tng nng estrogen,

progesterone trong huyt tng, ng thi lm gim prolactin huyt tng; c

tc dng gim cc triu chng tin mn kinh [17]. Tip n l nghin cu trn

loi Vitex rotundifolia L. T dch chit cn ca qu Vitex rotudinfolia L. thu

c 4 thnh phn chnh l: casticin,luteolin, rotundifuran v agnuside

c th nghim v cho hot ng ging nh estrogen[23]. Cc nghin cu

v thnh phn ha hc ca qu Vitex trifolia L. (mn kinh t) chng minh

n c cha cc thnh phn: casticin, luteolin (flavonoid ), rotundifuran

36

(diterpen), vitexilactone (lactone) v c bit n l cc cht c cu trc

phytoestrogen. Cc cht ny cng c phn lp t qu ca Vitex agnus

catus L., Vitex rotundifolia L. v cho tc dng nh estrogen. T nhng bng

chng trong kinh nghim y hc dn gian v cc bng chng thc nghim ca

y hc hin i,chng ti d on qu ca Vitex trifolia L. (mn kinh t) c

th c tc dng ph n giai on mn kinh v tin hnh nghin cu tc

dng gim lo lng, ci thin tr nh ca mn kinh t trn m hnh OVX

Trong nghin cu ny chng ti s dng m hnh OVX l m hnh

ng vt thc nghim nh gi tc dng hng estrogen, ci thin tr nh /

nhn thc, gim lo lng, chng long xng, nh gi trn than nhit, tim

mch, h hp ph n giai on mn kinh. M hnh ny hin ang c s

dng ph bin trn th gii tuy nhin cha c nghin cu no p dng m

hnh OVX nh gi tc dng gim lo lng, ci thin tr nh ph n mn

kinh v cng cha c nghin cu v tc dng ca Mn kinh t ph n giai

on mn kinh.

Kt qu thc nghim cho thy cao chit cn mn kinh t liu 3,2g/kg

c tc dng lm gim kh nng vn ng v ci thin tr nh chut OVX

bng th nghim nh gi hot ng t nhin v th nghim m l nc

Morris. Mc d trn th gii c mt s nghin cu v thnh phn ha hc

v tc dng sinh hc ca mn kinh t, nhng kt qu nghin cu ca chng

ti l hon ton mi v hin cha c kt qu nghin cu no v tc dng gim

lo lng, ci thin tr nh ph n mn kinh ca mn kinh t c cng b.

y l nhng kt qu ban u v l tin cho cc nghin cu su hn v

thnh phn ha hc cng nh tc dng hng estrogen ca mn kinh t.

37

KT LUN V XUT

1. KT LUN

Trong khun kh kha lun tt nghip vi ti Nghin cu thnh

phn ha hc v tc dng ci thin tr nh, gim lo lng ca cy Mn kinh

t Vitex trifolia L thu c cc kt qu sau:

V c im vi hc: Kt qu soi c im bt dc liu ( mn kinh t ) v

vi phu ca qu Mn kinh ph hp vi c im c m t trong DVN IV.

V thnh phn ha hc: nh tnh cc nhm cht hu c bng phn ng

ha hc v sc k lp mng xc nh c thnh phn ca Mn kinh t

gm c tinh du ( pht hin 10 vt trn sc k ), flavonoid ( pht hin 3 vt

trn sc k ), alkanoid, acid hu c.

V tc dng dc l: cao chit cn mn kinh t liu 3,2g/kg c tc

dng lm gim kh nng vn ng v ci thin tr nh/nhn thc chut

OVX bng th nghim nh gi hot ng t nhin v th nghim m l

nc Morris. Ngoi ra cn c tc dng tng trng lng t cung bung

trng.

2. XUT

Do thi gian c hn, kt qu nghin cu ca chng ti mi ch l nhng

bc u, chng ti xut:

Nghin cu su hn v thnh phn ha hc ca mn kinh t. nh lng

cc thnh phn c tc dng dc l nh tinh du, flavonoid. Phn lp cc

thnh phn cc dung mi khc nhau.

Tip tc nghin cu tc dng ca cao chit cn mn kinh t cc liu khc

v nghin cu tc dng trn cc phn on chit khc nhau.

Nghin cu cc tc dng dc l khc nh: tc dng trn t bo ung th,

tc dng khng Histamin.

38

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