Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6

KHO ST HOT TNH KHNG KHUN V KHNG OXY HA CA

CAO METHANOL CY H TH TRNG (Streptocaulon juventas MERR.)

i Th Xun Trang1, Lm Hng Bo Ngc1 v V Th T Anh1

1
Khoa Khoa hc T nhin, Trng i hc Cn Th

Thng tin chung:
Ngy nhn: 07/05/2015
Ngy chp nhn: 27/10/2015
Title:
Studies on antibacterial and
antioxidant activities of
methanolic extract from
Streptocaulon juventas Merr.
T kha:
Cy H Th , cht khng
oxy ha, DPPH, khng khun
Keywords:
Antioxidant, anti-bacteria,
DPPH, Streptocaulon
juventas
ABSTRACT
The purpose of this research is to study antimicrobial and antioxidant
activities of Streptocaulon juventas (Merr.). Antimicrobial and antioxidant
activities of vine and leaf methanol extracts of S. juventas were tested
using the Kirby-Bauer and DPPH methods, respectively. The results
proved that the methanol extract of S. juventas had high efficiency in
antimicrobial activity with the MIC value at16 g/ml on two bacterial
strains Escherichia coli and Staphylococcus aureus and higher
antimicrobial rate than that of the antibiotic ampicillin (MIC = 64 g/ml)
and amoxicillin (MIC E. coli = 64 g/ml and MIC S. aureus = 16 g/ml). The
results also showed that the DPPH scavenging activity of S. juventas was
high (IC50 = 349.35 g/ml). However, the antioxidant ability was 15.5
times lower than that of vitamin C (IC50 = 22.55 g/ml).
TM TT
Kh nng khng khun v khng oxy ha ca cao methanol cy H Th
trng (thn v l) c kho st. B phn ca cy H Th c ly
trch bng dung mi methanol. Kh nng khng vi khun Escherichia coli
v Staphylococcus aureus c xc nh bng phng php Kirby-Bauer
(Bauer et al., 1959; Bauer et al., 1966; Hudzicki, 2014) v kh nng khng
oxy ha c tin hnh bng phng php s dng DPPH (2,2-diphenyl-1
picrylhydrazyl). Kt qu chng minh hiu qu khng khun ca cao
methanol cy H Th rt cao gi tr MIC =16 g/ml i vi 2 dng vi
khun Escherichia coli v Staphylococcus aureus. Cao chit methanol cy
H Th c kh nng khng vi khun Escherichia coli v Staphylococcus
aureus cao hn thuc khng sinh chun ampicillin (MIC = 64 g/ml) v
amoxicillin (MICE. coli = 64 g/ml v MICS. aureus = 16 g/ml). Hiu qu loi
b gc t do hydro DPPH ca cao H Th (IC50 = 349,35 g/ml) thp
hn so vi vitamin C 15,5 ln (IC50 = 22,55 g/ml).

1 GII THIU gy bnh nhiu c quan khc nh tit niu, thn

kinh, h hp chng gy ra cc chng vim
Hin nay, nhim khun ng tiu ha tr nn
ng tit niu, nhim khun huyt. C th ni
rt ph bin v c th gy ra nhiu bnh nguy him
khi qut bt k bnh phm no cng c th gp
c bit i vi tr nh v c th gy t vong. Vi
thnh vin ca h vi khun ng rut. Ngy nay,
khun ng rut c chng minh l nguyn
phng php cha tr ch yu l s dng khng
nhn chnh gy tiu chy ngi. Ngoi ng
sinh nhng vic ny c th gy ra nhiu ri ro do
tiu ha, cc vi khun ng rut cn c kh nng

1
Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6
hin tng khng thuc ( Th Ty Phng,
2007). Bn cnh , stress oxy ha ang l mi
quan tm hng u vi cc nh khoa hc hin nay.
Stress oxy ha l hin tng xut hin trong c th
sinh vt khi c s mt cn bng gia vic sn xut
cc gc t do v hot ng ca cc cht khng oxy
ha. Hin tng ny l nguyn nhn ca rt nhiu
bnh nguy him trong c ung th cc bnh tim
mch, cc bnh suy gim h thn kinh (Alzheimer,
Parkinson) v lo ha sm (Li Th Ngc H v
V Th Th, 2009).
Cy H Th trng (Streptocaulon juventas
Merr.) v cc b phn ca cy c cc nh khoa
hc quan tm trong nhng nm gn y. Cc th
nghim v hot tnh khng ung th cho thy dch
chit methanol r cy H Th trng c c tnh
chn lc i vi nm dng t bo ung th l t bo
ung th t cung Hela ngi, t bo ung th phi
ngi A549, t bo ung th chut colon 26-L5, t
bo ung th phi chut LLC v t bo ung th rut
kt chut B16-BL6 chut (Ueda et al., 2002). Dch
chit ca H Th (c cy) c kh nng khng
khun (Quang-Vinh and Jong-Ban, 2013) v khng
oxy ha (Quang-Vinh and Jong-Ban, 2011). Trong
nghin cu ny, thnh phn s dng ca cy H
Th l thn v l vi mc ch kho st kh nng
khng khun v khng oxy ha.
2 PHNG PHP NGHIN CU
2.1 Phng tin
Thit b c s dng trong nghin cu gm
my c quay chn khng Heidolph (c), my o
pH Metler Toledo, my o quang ph, t cy
(Class II BSC, Esco, Indonesia).
Ha cht s dng trong th nghim gm:
methanol (Merck), mi trng Luria
Bertani (LB), DPPH (2,2 Diphenyl 1
picrylhydrazyl) (Wako, Japan), viatmin C, khng
sinh thng mi c s dng nh cht khng sinh
chun gm amipicillin (500 mg) v amoxicillin
(500 mg).
Vt liu th nghim l cy H Th trng
(Streptocaulon juventas Merr.) c thu hi Bn
Tre. Cy H Th sau khi thu mu c em v
phng th nghim v nh danh theo Phm Hong
H (2003).
Cc chng vi khun c s dng trong
th nghim bao gm vi khun Escherichia
coli, Staphylococcus aureus v Pseudomonas
aeruginosa do Trung tm kim nghim thuc - m
phm - dc phm Cn Th cung cp. Nhng
chng vi khun c chn l cc dng c s
dng rng ri trong cc nghin cu v tnh khng
khun, 3 chng vi khun ny c kh nng gy bnh
nghim trng v ng tiu ha trn ngi v cc
loi ng vt.
2.2 Phng php
2.2.1 iu ch cao methanol cy H Th
Bt th thn v l cy H Th (500 g) c
ngm trong methanol 48 gi, sau hn hp c
c quay p sut thp, thu c cao methanol
dng st.
2.2.2 Kho st s khng khun ca cao
methanol H Th
Chun b cao chit: cao chit c pha vi
dung mi methanol thnh cc nng 8, 16, 32,
64, 128 g/ml. Dch nui vi khun c pha long
trong nc mui sinh l tng ng c
0,5 Mc Farland mt s vi khun l 108 c tri
u trn mi trng LB c. a thch c
kh 15 pht trc khi t khoanh giy c tm cao
H Th .
Cao chit H Th cc nng kho st
khc nhau (100 l) c cho ln khoanh giy
(ng knh 6 mm) v trng. Cc khng sinh
ampicillin v amoxicillin c s dng nh i
chng dng v c pha thnh cc nng tng
t nh cao chit. Ngoi ra, do s dng methanol
pha cao chit nn nh hng ca methanol ln s
pht trin ca vi khun cng c kho st. Mi
a thch c t t 1, 2 hay 3 khoanh giy tm
cao chit, sau kh. Cc nng 8, 16, 32, 64,
128 g/ml c s dng trong kho st, mi nng
c lp li 3 ln. Cc a thch c 32C
trong 24 - 48 gi. ng knh vng c ch c
o bng thc o n v mm.
2.2.3 Kho st kh nng khng oxy ha DPPH
Kh nng khng oxy ha ca cao chit
methanol cy H Th trng c thc hin theo
phng php DPPH (2,2 Diphenyl 1
picrylhydrazyl) nh sau: cao methanol H Th
c pha thnh cc nng 100, 200, 300, 400,
500 g/ml trong methanol. Lng cao chit c
pha vo phn ng l 100 l v DPPH 610-4 M l
100 l (mi nng lp li 3 ln). Hn hp phn
ng c trong 60 pht trong ti, sau c o
hp thu quang ph bc sng 517 nm. Kh
nng khng oxy ha c tnh da vo hiu sut
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Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6

phn ng v hm lng cht khng oxy ha tnh 3.2 Kho st kh nng khng khun ca
tng ng vitamin C. cao methanol cy H Th
2.2.4 Thng k phn tch s liu Kh nng khng khun ca cao H Th c
Kt qu c x l thng k theo phng php xc nh da trn kh nng c ch s pht trin ca
phn tch Anova bng phn mm Minitab 16.0 v vi khun th hin qua ng knh vng khng
th c biu din bng phn mm Microsoft khun c to ra trn a petri c trnh by
Excel. Hnh 1 v Hnh 2. Kt qu m t Hnh 1 cho thy
cao H Th c kh nng khng khun i vi
3 KT QU V THO LUN hai dng vi khun Escherichia coli v
3.1 iu ch cao methanol cy H Th Staphylococcus aureus, ng knh vng v khun
c to ra nng cao kho st rt thp (< 16
Cao methanol cy H Th sau khi c quay g/ml). Ngc li, cao H Th khng c kh
thu c 20,04 g cao dng st vi hiu sut chit nng khng vi khun Pseudomonas aeruginosa
cao l 3,55% (so vi trng lng kh). (vn c vng v khun nhng khun lc pht trin
khp vng Hnh 1C).

A B C

Hnh 1: Vng v khun ca cao vi cc dng vi khun (mm)

(A): Escherichia coli (B): Staphylococcus aureus (C): Pseudomonas aeruginosa

Xt mt cch tng th v kh nng khng E.coli cao hn vi khun S. aureus mt cch khc
khun ca cao methanol H Th i vi hai bit c ngha thng k nng kho st t 16
dng vi khun E. coli v S. aureus cho thy cao H g/ml n 128 g/ml (Hnh 2).
Th c kh nng c ch s pht trin vi khun

35
ng knh vng v khun (mm)

30

25

20
E. coli
15
S. aureus
10

0
0 20 40 60 80 100 120 140
Nng cao H Th (g/ml)
Hnh 2: Kh nng khng khun ca cao H Th trn 2 loi vi khun

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Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6

Hiu qu khng khun ca cao H Th i ampicillin. nng 16 g/ml, cao H Th c

vi chng vi khun Escherichia coli v hot tnh c ch cao vi 2 chng vi khun E. coli
Staphylococcus aureus trnh by Bng 1. Kt qu v S. aureus to vng v khun r rt, ln lt l
cho thy ng knh vng khng khun t l thun 25,3 0,6 mm v 22,7 0,6 mm (Bng 1), trong
vi nng cao H Th ; ngha l kh nng khi khng sinh ampicillin khng c ch hai
khng khun ca cao H Th tng khi tng nng dng vi khun ny nng 16 g/ml (Hnh 3).
cao chit. Hiu qu khng khun ca cao H S nhy cm ca vi khun vi khng sinh l rt
Th i vi chng vi khun E. coli cao quan trng iu tr cc bnh nhim khun.
hn khc bit c ngha thng k so vi vi Trong nghin cu ny cho thy hai dng vi khun
khun S. aureus tt c cc nng kho st kho st u nhy cm vi cao H Th , iu ny
(Bng 1). So snh v nhy cm ca vi khun vi chng minh rng cc tho dc c th thay th
khng sinh th vi khun E. coli v S. aureus nhy cc khng sinh thng mi nhng mc an ton
cm vi cao H Th hn so vi khng sinh cao hn.
Bng 1: Kh nng khng khun ca cao methanol cy H Th
Nng cao ng knh vng v khun d SE (mm)
(g/ml) E. coli S. aureus P. aeruginosa
0 - - -
8 10,3d 0,6 11,7c 1,5 -
16 25,3c* 0,6 22,7b 0,6 -
32 26,7bc* 0,6 23,3ab 0,6 -
64 27,7ab* 0,6 24,0ab 1,0 -
128 29,0a* 1,0 25,3a 0,6 -
Ghi ch: kt qu vi lch chun ca tng gi tr. Cc mu t theo sau cc gi tr trong cng mt ct khc nhau th
khc bit c ngha thng k mc 5%. (*) l s khc bit gia hai dng vi khun E.coli v S. aureus c ngha mc
5% tng nng cao chit c kho st. (-) l khng khng khun
So snh kh nng khng khun ca cao kho st. Biu trnh by Hnh 3B cho thy,
methanol H Th vi thuc khng sinh thng kh nng khng vi khun S. aureus ca cao H Th
mi c s dng l ampicillin v amoxicillin cao hn khng sinh ampicillin tt c nng
c trnh by Hnh 3. Biu trnh by Hnh kho st. Tuy nhin, cao H Th c kh nng
3A cho thy, kh nng khng vi khun E. coli ca khng vi khun S. aureus thp hn khng sinh
cao H Th cao hn so vi hai loi khng sinh amoxcillin khi kho st nng t 32 g/ml n
chun amipicillin v amoxicillin tt c nng 128 g/ml.

A B
Hnh 3: Kh nng khng khun ca cao H Th v hai loi khng sinh
(A): vi khun E.coli, (B): vi khun S. aureus
Kh nng khng khun ca cao H Th c khun ca cao H Th vi hai loi khng sinh
xc nh da trn gi tr MIC (minimum inhibitory ampicillin v amoxicillin c trnh by Bng 2.
concentration). Kt qu so snh kh nng khng

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Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6
Bng 2: Nng c ch vi khun ti thiu
(MIC) ca cao v 2 loi khng sinh
MIC (g/ml)
S.
E. coli
aureus aeruginosa
Cao HTO 16 16
Ampicillin 64 64
Amoxicillin 64 16
(-): khng khng khun
Nng c ch ti thiu l nng cao chit
(hoc khng sinh) thp nht m ti xut hin
vng v khun; nn nng c ch ti thiu cng
thp th kh nng khng khun cng cao. Kt qu
trnh by Bng 2 cho thy, nng c ch ti
thiu ca cao H Th hai chng vi khun E.
coli v S. aureus bng nhau v bng 16 g/ml, cao
H Th khng c tc dng khng vi khun P.
aeruginosa. Kh nng khng hai chng vi khun
E. coli v S. aureus ca ampicillin (MIC =
64 g/ml) thp hn so vi cao H Th
(MIC = 16 g/ml). Khng sinh ampicillin c kh
nng khng P. aeruginosa (MIC = 64 g/ml), trong
khi cao H Th khng c kh nng khng chng
vi khun ny. Khng sinh amoxicillin c kh nng
khng khun trn ba dng vi khun kho st l E.
coli, S. aureus v P.aeruginosa vi gi tr MIC ln
lt l 64 g/ml, 16 g/ml v 64 g/ml. Kt qu
cho thy, kh nng khng vi khun E. coli v S.
aureus ca cao H Th (MIC = 16 g/ml) cao
hn khng sinh amoxicillin. Kt qu c th chng
minh kh nng khng khun ca cao H Th cao
hn gp 4 ln so vi khng sinh ampicillin trn hai
dng vi khun E. coli v S. aureus. Cao methanol
cy H Th c hiu qu khng vi khun E. coli
cao gp 4 ln so vi khng sinh amoxicillin. Hiu
qu khng vi khun S. aureus ca cao H Th
tng ng vi khng sinh amoxicillin.
Hot tnh khng khun ca cao H Th c
so snh vi cc loi cao chit khc trn cng ba
dng vi khun kho st thng qua gi tr MIC
(g/ml). Cao H Th (MIC = 16 g/ml trn c
hai dng vi khun E. coli v S. aureus) c kh nng
khng vi khun E. coli v S. aureus cao hn
so vi thn Chiu liu ngh (Terminalia
nigrovenulosa) vi MIC ln lt l 625 g/ml v
156 g/ml. Kh nng khng khun ca cao H
Th cng cao hn so vi cao l Chiu liu
ngh (Terminalia nigrovenulosa) trn vi
khun E. coli (MIC = 312 g/ml) v vi khun
S. aureus (MIC = 78 g/ml). Hot tnh khng
khun cao H Th (MIC = 16 g/ml trn c hai
dng vi khun E. coli v S. aureus) cng cao hn
so vi Premna integrifolia L. (c cy) trn vi
khun E. coli (MIC = 625 g/ml) v vi khun S.
aureus (MIC = 625 g/ml) (Quang-Vinh and Jong-
P. Ban, 2013). Kh nng khng khun ca cao H
Th (thn v l) trong kho st cao hn so vi
cao cy H Th (c cy) dng so snh trn vi
-
khun E. coli (MIC = 2500 g/ml) v trn vi khun
64
S. aureus (MIC = 156 g/ml) (Quang-Vinh and
64
Jong-Ban, 2013). Tuy nhin, cc loi cao chit
c so snh vi cao H Th u c kh nng
khng vi khun P. aeruginosa (MIC 1250 g/ml)
(Quang-Vinh and Jong-Ban, 2013), trong khi
cao H Th (thn v l) khng c tc dng
khng vi khun ny trong cc nng kho st.
3.3 Kho st kh nng lm sch gc t do
DPPH ca cao chit cy H Th
DPPH l phng php c s dng rng ri
kim tra kh nng loi b gc t do v cc
nhm cho hydro, phng php ny cng c s
dng nh lng cc cht oxy ha trong h
thng sinh hc phc tp ngy nay. Nhng electron
l c trong gc t do DPPH cho s hp thu mnh
nht bc sng 517 nm v hp cht ny c mu
tm. Khi cc electron l ny kt hp vi hydro ca
cht khng oxy ha hnh thnh dng DPPH-H,
hp cht s chuyn t mu tm sang vng tng
ng vi lng electron kt hp vi DPPH (Prakash
et al., 2000). V vy, kh nng lm sch gc t do
ca mt cht cng cao th s hp thu quang ph
c o bc sng 517 nm ca phn ng DPPH
c gi tr cng thp v ngc li.
Kh nng khng oxy ha ca cao H Th
c kho st da trn hm lng cht khng oxy
ha. Hm lng cht khng oxy ha c tnh
tng ng g/ml vitamin C da vo phng
trnh ng chun y = -0,029x + 1,3311 (R2 =
0,9897).
Kt qu khng oxy ha ca cao methanol cy
H Th c trnh by trong Bng 3 cho thy,
hiu sut khng oxy ha ca cao H Th t l
thun vi nng cao v khc bit c ngha
thng k cc nng kho st. Hiu sut khng
oxy ha ca cao H Th cao nht l 72,07%
nng 500 g/ml.
Kt qu hm lng cht khng oxy ha c
trong cao H Th c tnh da vo phng
trnh ng chun viatmin C (y = -0,029x +
1,3311) c trnh by Bng 3. Kt qu trnh by
Bng 3 cho thy, nng cao t 0 n 500 g/ml
c lng vitamin C tng ng tng dn t 0 n
33,15 0,86 g/ml. Nng cao methanol H Th
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Tap ch Khoa hoc Trng ai hoc C n Th Phn A: Khoa hc T nhin, Cng ngh v Mi trng: 40 (2015): 1-6

cao nht 500 g/ml tng ng vi 33,15 0,86 = 349 g/ml thp hn vitamin C (IC50 = 22,55
g/ml vitamin C v 100 g/ml l nng thp nht g/ml) 15,5 ln.
vi lng vitamin C tng ng 9,8 0,52 g/ml.
TI LIU THAM KHO
Bng 3: Hiu sut khng oxy ha ca cao H
Phm Hong H, 2003. Cy c Vit Nam. Nxb
Th (thn v l)
Tr, TP. H Ch Minh.
Nng cht i Th Xun Trang, Nguyn Th Mai Phng,
Nng Hiu sut
khng oxy ha V Th Ngc Dim v Quch T Hu, 2012.
cao khng
tng ng Kho st hiu qu h ng huyt v chng
(g/ml) oxy ha (%)
vitamin C (g/ml) oxy ha ca cao chit cy Nhu (Morindan
0 0 0 citrifolia L.) chut bnh tiu ng. Tp ch
100 20,94e 0,45 9,80e 0,52 Khoa hc, i hc Cn Th: 23b; 115-124.
200 37,07d 1,31 17,16d 0,43
300 50,57c 2,13 23,33c 0,97 Th Ty Phng, 2007. Xy dng quy trnh
400 63,39b 1,04 29,18b 0,69 k thut tch chit, kho st tnh khng
500 72,07a 2,25 33,15a 0,86 khun v kh nng chng oxy ha ca mt
s hp cht th cp t l cy Xun Hoa
Ghi ch: cc mu t theo sau cc gi tr trong cng mt (Pseudranthemum palatiferum). Kha lun
ct khc nhau th khc bit ngha v mt thng k
tt nghip. i hc Nng Lm TP. H Ch
mc 5%
Minh: 20 23.
Hiu qu loi b gc t do ca cao methanol Li Th Ngc H v V Th Th, 2009. Stress
cy H Th c xc nh thng qua gi tr IC50 oxy ha v cc cht khng oxy ha t nhin,
c trnh by Bng 4, kh nng khng oxy ha Tp ch Khoa hc v Pht trin, 667-677.
ca cao H Th (IC50 = 349,35 g/ml) thp hn
Bauer AW, Perry DM and Kirby WMM, 1959.
so vi vitamin C (IC50 = 22,55 g/ml) c dng
Single disck antibiotic sensitivity testing of
kho st trong th nghim 15,5 ln. Kt qu
Staphylococci. A.M.A. Arch. Intern. Med.
nghin cu ny chng minh cao H Th c kh
104:208216.
nng khng oxy ha cao hn mt s tho dc
trong cc nghin cu trc y nh cy Nhu Bauer AW, Kirby WMM, Sherris JC and Turck
(Morinda citrifolia L.) gm cao ethanol l, tri M, 1966. Antibiotic susceptibility testing by
xanh, r cy Nhu vi gi tr IC50 ln lt l 917,16 a standardized single disk method. Am. J.
g/ml, 1025,2 g/ml v 1531,4 g/ml (i Th Clin. Pathol. 36:493-496.
Xun Trang v ctv, 2012); cy C gai leo (Solanum Hudzicki J, 2014. Kirby-Bauer Disk Diffusion
hainanense Hance) IC50 l 1734 g/ml v cao H Susceptibility Test Protocol. American
Th (Streptocaulon juventas Merr.) (c cy) c Society for Microbiology MicrobeLibrary.
kh nng khng oxy ha vi gi tr l IC50 = 2586 Prakash A, Rigelhof F, and Miller E, 2000.
g/ml (Quang- Vinh and Jong-Ban, 2011). Antioxidant activity. Analytical progress
Bng 4: Gi tr IC50 ca cao methanol HTO so Medallion Laboratories, 1-4.
vi vitamin C Quang-Vinh Nguyen, Jong-Ban Eun, 2011.
Gi tr IC50 (g/ml) Antioxidant activity of solvent extracts
Cao HTO 349,35 from Vietnamese medicinal plants. Journal
Vitamin C 22,55 of Medicinal Plants Research Vol. 5(13),
2798-2811.
4 KT LUN Quang-Vinh Nguyen, Jong-Ban Eun, 2013.
Cao methanol H Th trng (thn v l) c Antimicrobial activity of some Vietnamese
kh nng khng hai loi vi khun Escherichia coli medicinal plants extracts. Journal of
v Staphylococcus aureus cao hn thuc khng Medicinal Plants Research Vol. 7(35), pp.
sinh thng mi ampicillin v amoxicillin. Cao 2597-2605.
methanol H Th trng khng c kh nng Ueda JY, Tezuka Y, Banskota AH, Tran QL,
khng vi khun Pseudomonas aeruginosa cc Tran QK, Harimaya Y, Saiki I, Kadota S,
nng kho st. 2002. Antiproliferative activity of
Vietnamese medicinal plants. Biol. Pharm.
Cao methanol H Th c kh nng khng
Bull. 25:753-760.
oxy ha trong phng php DPPH vi gi tr IC50