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Isolation and Extraction of Total Flavonoids From Epimedium Koreanum Nakai by Supercritical Fluid Extraction PDF
Isolation and Extraction of Total Flavonoids From Epimedium Koreanum Nakai by Supercritical Fluid Extraction PDF
Coll ege
,
Changchun
1332,
P
.
R
.
China
!
3.
China
-
Japan nion ! ospital
,
Jili n ni" ersity
,
Changchun
1332,
P
.
R
.
China
"e#ei$ed June
1, 2%
* &upp'( )ed *+ )he Na)u(al &#ien#e and , e#hn'l'g+ -'unda)i'n '. Jilin /( '$in#e0 N'. 22132-13 .
* * ,' 4 h'm #'( (e5p'nden#e 5h'uld *e add( e55ed. 6-mail7 liu8q9 #ia#. :l. #n
I n )he p( e5en) pape( i5 ( ep'( )ed )he me)h'd .'( )he i5'la)i'n and e;)(a#) i'n '. )') al .la$'n'id5 '.
#pimedium $ or eanum N a%ai *+ mean5 '. 5upe(#( i)i#al .luid e;) (a#)i'n0 &-63 . <+ e;amining p(e55u(e, )em-
pe(a) u( e
,
am'un)5 '. m'di.ie( and e;) (a#)i'n )ime
,
)he 'p)imi8ed #'ndi)i'n '. &-6 i5 #'n.i( med a5
3
=/a
and 1 , 4 i)h 2> e)han'l a5 )he m'di.ie( . , he 5ample5 4 e(e 5)a)i#all+ e;)( a#) ed .'( 3 min, .'ll'4 ed *+
d+nami# e;)( a#)i'n .'(
12
min a) a .l'4 (a)e '.
1
mL
?
min
.
,he quan)i)a)i$e anal+5i5 '. )')al .la$'n'id5 4 a5
pe( .'( med *+ U@-@i5 5pe#) ('ph')'me)( +. C'mpa(ed 4i)h ) he #'n$en) i'nal me)h'd, )he &-6 me)h'd i5 m'( e
e..i#ien), m'(e ( apid and m'( e .( iendl+ en$i('nmen)all+.
Keywords &upe( #( i)i#al .luid e;)( a#)i'n0 &-63 , -la$'n'id5, #p imedium $ or eanum N a%a
Articl e ID 1A-B%0 2%3 -1-22-%
Introduction
,he he(*al pl an) #pimedi um $oreanum Na%ai
0 &er ber idaceae3 i5 a na) i$e he(* g('4ing in ) he
Chang*ai ='un) ain (egi'n, Jilin p('$in#e '. China,
and i5 u5ed a5 a ) 'ni#, an) i-(heuma) i# and aph('-
di5ia# in ) (adi) i'nal Chine5e medi#ine, ) 'ge)he(
4i)h . 'u( ') he( Chine5e /ha(ma#'p'#ia-(e#'(ded
5pe#ie57 6pimedium *(e$i#'(num =a;im, 6pimedi-
um 5agi)) a) um =a;im, 6pimedium pu*e5#ene5
=a;im and 6pimedium 4u5hanen5e , . &. Cing
D 1E
.
I) ha5 *een (ep'() ed ) ha) in #pimedium $ ore-
anum Na%ai ) he(e a( e )he #'mp'nen)5
.l a$'n'id5
D 2 1E
,
alFal'id5
D 11 13E
,
p'l+5a##ha-
(ide5
D 1%, 1AE
, mi#('elemen) 5
D 11, 12E
, and 5' 'n, am'ng
4hi#h . la$'n'id5 a(e ) he main #'mp'nen) 5. "e#en)
pha(ma#'l'gi#al 5) udie5 dem'n5) (a) e ) ha) ) ') al
.l a$'n'id5 in #p imedium $oreanum Na%ai 5h'4
) he an)i-in.lamma) '(+ e. .e#) 5
D 1GE
and '*$i'u5 h+-
p') en5i$e a#) i$i)+
D 1BE
. , he+ #an imp( '$e )he im-
mune . un#) i'n
D 2E
, in#(ea5e )he *'ne den5i) +, en-
han#e ) he e5) (adi'l le$el , de#( ea5e IL-1 #'n#en) (a-
)i'n in 5e(um, and p(e$en) '5) e'p'( '5i5
D 21 23E
.
Up ) ill n'4, )')al .la$'n'id5 ha$e *een mainl +
i5'l a) ed .('m #pimedi um $ oreanum Na%ai *+ e;-
)( a#) i'n 4i) h e) han'l
D 2%E
. In ) hi5 ) ime-#'n5umed
p( '#e55, la(ge am'un) 5 '. 5'l$en) 5 a(e #'n5umed
4hi#h ma+ p'llu)e )he en$i('nmen) *adl +.
Cu( (en) l+, )he &-6 ) e#hnique a. .'(d5 man+
ad$an) age5 '$e( )he m'(e #'n$en) i'nal e;) (a#)i'n
me) h'd. &upe(#(i) i#al #a(*'n di';ide i5 4idel+ u5ed
a5 an e;) (a#) i'n 5'l$en) due ) ' i) 5 l'4 #(i) i#al
p'in) , n'n-)';i#i) +, #hemi#al ine() ne55, n'n-e;pl'-
5i$ene55, l'4 $ap'( l'55, a$aila*ili)+ and ea5e '.
(em'$al . ('m ) he e;) (a#)ed 5+5) em5. -u() he(m'(e,
) he e;)( a#) d'e5 n') need ) ' *e #'n#en) (a)ed *e-
#au5e 'nl+ a li) ) le am'un) '. '(gani# 5'l$en) i5
u5ed. , he5e ad$an) age5 ha$e a) ) (a#) ed in#(ea5ing
in) e(e5) .( 'm (e5ea(#he(5
.
Al) h'ugh &-6 ha5 *een
empl '+ed .'( ) he e;) (a#) i'n '. man+ #la55e5 '.
he( *5 and na) u(al p('du#)5
D 2A 33E
,
'nl + a .e4 5) ud-
ie5 in$'l$ing ) he 5epa(a) i'n '. .la$'n'id5 ha$e *een
pu*li5hed
D 3% 32E
. , ' ) he *e5) '. 'u( Fn'4ledge, n'
(ep'() ha5 +e) appea(ed 'n ) he &-6 '. #p imedium
$ oreanum Na%ai
.
In ) hi5 4'(F
,
4e . '#u5ed 'u( a) ) en) i'n 'n ) he
e;) (a#)i'n '. ) ') al . la$'n'id5 in #p imedium $ore-
anum Na%ai *+ &-6
.
,he in. luen#e5 '. p(e55u(e
,
) empe(a) u( e
,
am'un) '. e)han'l a5 ) he m'di. ie( and
e;) (a#)i'n ) ime 4e(e e$alua) ed in '( de( ) ' ge) 5a) i5-
.a#)'(+ (e5ul) 5
.
,he ) ') al .la$'n'id5 e;) (a#) ed 4e(e
anal+8ed *+ U@-@i5 5pe#) ('me)(+, and )he (e5ul) 5
4e( e #'mpa( ed 4i) h ) h'5e '*)ained *+ ) he #'n$en-
CH6=
.
"6&
.
CHIN6&6 U
. 2%, 20 13 , 22 21
)i'nal e;) (a#) i'n me) h'd
.
Experimental
1 Plant Materi al
,he lea$e5 '. #p imedium $ or eanum Na%ai
4e( e #'l le#) ed . ('m ) he Chang*ai ='un) ain, Jilin
/('$in#e '. China in Jul+, 23, and ) he ma;imal
am'un) '. . la$'n'id5 4a5 p('du#ed and u5ed .'( all
) he e;pe(imen) 5. , he plan) ma) e(ial 4a5 d(ied in
) he 5hade . '( 5e$e(al da+5 and ) hen pul$e(i8ed.
2
Equi pment and Reagents
&-6 4a5 pe(.'(med 'n an
0
HA
221-A-13
5u-
pe(#(i)i#al .luid e;) (a#) '(
0
Nan) 'ng
,
Huaan =a-
#hine -a#) '(+
3 ,
4hi#h i5 de5igned . '( 4'(Fing
p( e55u(e5 and )empe(a)u(e5 up ) '
A
=/a and
1 ,
(e5pe#)i$el+
.
A U@
-
@i5 5pe#)( 'ph') 'me) e(
0
='del
11,
Agilen)
3
4i)h
1
#m qua() 8 a*5'( p-
)i'n #ell5 4a5 u5ed ) ' anal+8e quan) i) a) i$el+ ) he ) '-
)al . la$'n'id5 a) 22 nm. Anal+) i#al g(ade e)han'l
4a5 u5ed a5 a m'di.ie( and . '( )he #'n$en) i'nal e;-
)( a#) i'n '. ) he ) ') al . la$'n'id5. Hi5) illed 4a) e( 4a5
u5ed in all ) he e;pe(imen) 5.
!onventional Extraction
,he d(ied and pul$e(i8ed pl an) ma) e(ial '.
A. g 4a5 e;) (a#) ed 4i) h A mL '. 2> e)han'l
unde( (e.l u; di5) illa) i'n . '( 2 h. , he e;) (a#) i'n
4a5 (epea) ed .'( ) h(ee ) ime5 in '(de( ) ' #'mple) el+
e;) (a#) ) he e. .e#) i$e #'mp'nen) 5 .( 'm ) he lea$e5 '.
#p imedium $ or eanum Na%ai
.
, he #'m*ined e;-
)( a#) 5 4e(e #'n#en) (a) ed in a (')a(+ e$ap'(a)'( a) a
(edu#ed p(e55u(e un) il a $i5#'u5 (e5idue (emained
.
,he (e5idue 4a5 e;) (a#) ed 4i) h pe)( 'leum e) he(
0
*
.
p
. 1 B 3
in '(de( ) ' (em'$e #hl'('ph+ll
,
and
) hen ) he aque'u5 la+e(5 4e(e e;) (a#) ed .'( ) h( ee
) ime5 4i)h e)h+l a#e) a) e
,
.inal l+
,
) he e) h+l a#e) a) e
la+e(5 4e(e #'m*ined and #'n#en) (a) ed a) a ( edu#ed
p( e55u(e ) ' +iel d 1 mL '. a +ell'4 5'lu) i'n.
" Supercritical Fluid Extraction
,he &-6 e;pe(imen) 4a5 pe(. '(med in ) he
p( e55u(e (ange .('m 2 ) ' 3A =/a and ) he ) empe(a-
)u(e ( ange .('m % ) ' 2 . , he 5upe(#(i) i#al . luid
.l '4 ( a) e 4a5 #'n) ('lled a) 1 mL? min. In ea#h e;-
pe(imen), ) he e;) (a#) '( 4a5 pa#Fed 4i) h A. g '.
d( + #pimedi um $oreanum Na%ai p'4de( , 4hi#h
4a5 mi;ed 4i) h A g '. gla55 *ead5 ) ' in#(ea5e ) he
#'n) a#) 5u(. a#e *e) 4een ) he plan) ma) e(ial and ) he
5upe(#(i) i#al #a(*'n di';ide unde( $a(i'u5 #'ndi-
)i'n5
.
<')h ) he ) 'p and *') ) 'm pa() 5 '. ) he #ell
4e( e . illed 4i)h a la+e( '. gl a55 4''l in '( de( ) ' de-
#(ea5e )he unu5ed 5pa#e
.
-'( all ) he m'di.ie( 5) ud-
ie5
,
) he m'di.ie(5 4e(e added *+ mean5 '. a 5epa-
(a) e pump 5' a5 ) ' ge) )he m'5) a##u(a#+ and (ep('-
du#i*ili) +. , he e;) (a#) ed anal+) e5 4e(e #'lle#) ed in
3 mL '. BA> e) han'l in a 1 mL . la5F a) (''m
) empe(a) u( e.
# T$e %nalysi s of Total Fl avonoids
,he am'un) 5 '. )he ) ') al . la$'n'id5 4e(e de-
) e(mined *+ mean5 '. U@-@i5 5pe#) ('me) (+ 4i) h
I#a((in a5 5) anda(d a##'( ding )' )he me)h'd .('m
(e.e(en#e
D 12E .
,he a*5'( *an#e 4a5 mea5u(ed a-
gain5) a *lanF a)
22
nm
,
and ) he )')al . la$'n'id5
pe(#en) age 4a5 e5)ima) ed 4i)h ) he aid '. ) he . 'l-
l'4ing linea( equa) i'n *a5ed 'n ) he #ali*( a)i'n
#u($e '. I#a(( in. A I . 3GB- . A1, r I
. BBB2, 4he( e A i5 ) he a*5'(*an#e $alue, and i5
) he I#a((in ma55 #'n#en) (a) i'n in g? mL. ,he #ali-
*( a) i'n #u( $e 4a5 linea( *e)4een 12 g? mL.
Results and &iscussion
1 T$e Inf luence of Modi fier on t$e Extraction Ef'
ficiency
<e#au5e '. ) he limi) ed 5'lu*ili)+ '. p'la( '(-
gani# #'mp'und5 in 5upe(#(i) i#al #a(*'n di';ide e;-
) (a#) i'n
,
)he quan) i) a) i$e e;) (a#) i'n '. p'la( ana-
l+) e5 4i) h pu(e 5upe(#(i)i#al .l uid '. CO
2
i5 di55a) i5-
.a#)'(+. ,' imp('$e ) he e;) (a#)i'n e. .i#ien#+ '. ) '-
) al . la$'n'id5, 2> e) han'l and 1> e)han'l
4e( e u5ed a5 )he m'di. ie(. 6)han'l 4a5 #h'5en due
) ' i) 5 l'4 ) ';i#i)+ #'mpa(ed 4i) h ) he ') he( '(gani#
5'l$en) 5. A) )he $e(+ *eginning '. d+nami# e;) (a#-
) i'n, ) he m'di. ie( 4a5 added *+ mean5 '. a 5epa-
(a) e pump in '(de( ) ' '*)ain ) he highe5) a##u(a#+
and (ep('du#i*il i) +
.
,he in. luen#e5 '. di. . e(en) $'l-
ume5 '. )he m'di. ie( 'n ) he e;) (a#) i'n e. .i#ien#+ '.
) ') al . la$'n'id5 a(e 5h'4n in ,a*le
1.
Table 1 T$e influence of modif ier on extracted
ef ficiency of total flavonoids
a
='di. ie(
Addi)i'n
am'un) 5? mL
6; )(a#) i'n
e. . i#ien#+
0 > 3
b
6) han'l
3 2. 1
A 2. G
2 3. %
2> 6) han'l 3 %. 1
A %. B
2 A. %
/u(e CO
2
1. 1
a. &upe(#(i) i#al #a(*'n di';ide a) 2A =/a, A ! b. mean $al-
ue5 0 nI 33 e;p(e55ed a5 ) he pe(#en) age (e. e(ed ) ' ) he d( + 4ei gh) .
,he e;pe(imen) al (e5ul) 5 dem'n5) (a) e ) ha) ) he
addi) i'n '. e) han'l #an 5u*5) an)iall+ enhan#e ) he
+ield '. ) ') al e;) (a#) ed . la$'n'id5. A) a #e() ain
) empe(a) u( e and a p(e55u( e, ) he highe( ) he am'un)
'. ) he m'di.ie( i5, ) he highe( ) he e;) (a#)i'n +iel d
2G
CH6=
.
"6&
.
CHIN6&6 U
.
@'l
. 2
a#hie$ed i5
.
<u) 4hen )he 5ame am'un) '. ) he
m'di.ie( i5 u5ed
,
)he e;)( a#) ed +ield5 4i) h
2>
e)han'l a5 ) he m'di. ie( a(e highe( ) han ) h'5e 4i) h
pu(e e) han'l a5 ) he m'di. ie(
.
, hi5 #'uld *e e;-
plained *+ ) he . a#) ) ha) p'la( #'n5)i) uen) 5 in ) he
pl an) 4'uld *e ea5ie( )' *e e;)( a#) ed 4i)h a m'(e
p'la( 5'l $en)
D 3AE
.
A5 a (e5ul )
, 2
mL '.
2>
e)han'l 4a5 5ele#)ed in ) he 5u*5equen) e;pe(i-
men) 5
.
2
T$e Influence of Pressure on t$e Extraction Ef'
ficiency
,he p( e55u(e and ) empe(a) u(e a(e ) he m'5) im-
p'( )an) ph+5i#al pa(ame)e(5 in &-6 e;pe(imen)
,
4hi#h #an a. .e#) ) he anal+) e 5'lu*ili) + '*$i'u5l+
.
-'( e;amining )he in.luen#e '. p(e55u(e, ) he +ield5
'. ) ') al e;)(a#) ed . la$'n'id5 a) di. .e(en) p(e55u(e5
4e( e in$e5) iga) ed a) % and 4i) h 2> e) han'l a5
) he m'di.ie(. ,he e;pe(imen) al (e5ul) 5 5h'4 ) ha)
) he +iel d '. ) ') al e;)( a#) ed . la$'n'id5 in#(ea5e5
4i)h )he in#(ea5e '. ) he p(e55u(e $a(+ing . ('m 2 ) '
3 =/a, and ) hen, )he e;) (a#) i'n +ield in#(ea5e5
5ligh) l+ 4hile ) he p(e55u(e i5 in#(ea5ed .('m 3 ) '
3A =/a0 5ee -ig. 13 , 4hi#h dem'n5) (a) e5 ) ha) ) he
in#(ea5e '. ) he den5i) + '. 5upe(#(i) i#al #a(*'n di';-
ide (e5ul) 5 in ) he in#(ea5e '. ) he e;) (a#) i'n +ield.
<u) high p(e55u(e (e5ul) 5 in g(ea) e( #'5) . '( ) he e;-
)( a#) i'n 5+5)em, a5 4ell a5 )he 5) (ingen) 'pe( a) i'n
de5ign and in#( ea5ed ene( g+ demand
D 22E
. , he(e. '(e,
3 =/a i5 ) he 'p)imal p( e55u(e . '( )he e;)( a#) i'n.
Fig
. 1
T$e eff ect of pressure on t$e extraction eff iciency
of total flavonoids at #' .
T$e Influence of Temperature on t$e Extraction
Efficiency
,he e. .e#) '. )empe(a)u(e 'n ) he e;)( a#) i'n '.
) ') al .la$'n'id5 . ('m #p imedium $ oreanum Na%ai
4a5 in$e5) iga) ed 'n Feeping all ') he( e;) (a#) i'n pa-
(ame) e(5 ) ' *e . i;ed. , he e;) (a#) i'n e. .i#ien#+ '.
) ') al . la$'n'ide5 *+ 5upe(#(i)i#al . luid #a(*'n di';-
ide a) .'u( di.. e(en) ) empe(a) u(e50 %, A, 1 and
2 3 i5 5h'4n in -ig. 2. ,he e;) (a#) i'n +ield in-
#(ea5e5 al'ng 4i) h ) he )empe(a)u(e #hanging .('m
%
)'
1 ,
*u) 4i) h ) he ) empe(a) u(e in#(ea5e
. ('m
1
) '
2 ,
)he e;) (a#)i'n +ield de#(ea5e5
5l igh)l+
.
,he5e (e5ul) 5 dem'n5) (a) e ) ha) ) he in-
#(ea5e '. ) he $ap'( p(e55u( e #au5ed *+ ) he in#(ea5e
'. ) empe( a) u(e appea(5 ) ' #'mpen5a) e . '( ) he de-
#(ea5e in 5'lu*ili)+ #au5ed *+ d('pping '. 5upe(#(i) -
i#al . luid den5i) +
,
(e5ul) ing in )he e;)(a#) i'n +iel d
) ' in#(ea5e 4i) h ) he ) empe(a) u(e in#( ea5e .('m
%
) '
1 ,
*u) 4hen ) he )empe( a)u(e #'n) inue5 ) '
in#(ea5e
,
) he ( edu#) i'n '. 5upe( #(i) i#al #a(*'n di';-
ide den5i) + *e#'me5 d'mina) e in )he e;) (a#) i'n p('-
#e55
D 2B, 3%E
.
, hu5 an e;) (a#) i'n ) empe( a) u(e '.
1
4a5 5ele#) ed . '( e;)(a#) ing ) he m'5) '. ) ') al
. la$'n'id5
.
Fig. 2 T$e ef fect of temperature on t$e extraction eff i'
ciency of total f lavonoids at
2#
MPa
.
"
T$e Infl uence of Extraction Time on t$e Ex'
traction Ef ficiency
Fig. T$e ef fect of time on extraction eff iciency of
total f lavonoids at
2#
MPa and
#' .
A. )e(
3
min '. #'n5) an) 5) a)i# e;) (a#)i'n
) ime
,
) he e. .e#) '. ) he d+nami# e;) (a#)i'n ) ime
. ('m
3
) '
1A
min 'n ) he e;)( a#) i'n e. .i#ien#+ 4a5
5) udied
.
,he e;pe(imen) al (e5ul )5 dem'n5)( a) e ) ha)
) he l'nge( ) he 5upe(#( i) i#al #a(*'n di';ide i5 in #'n-
) a#) 4i) h )he ma) (i;
,
) he highe( ) he am'un) '. ) ') al
. la$'n'id5 e;) (a#)ed i5
0
5ee -ig
. 33 ,
and ) he e;) (a#-
) i'n +iel d in#(ea5e5 5'me4ha) a. ) e( )he d+nami# e;-
) (a#) i'n '.
12
min
.
&' )he #'m*ina)i'n '.
3
min
'. 5) a) i# e;) (a#)i'n and 12 min '. d+nami# e;) (a#-
N' 2B
. 1
LIU Chun
-
ming et al
.
)i'n 4a5 #h'5en . '( e;) (a#) i'n
.
A##'(ding ) ' )he (e5ul) 5 men) i'ned in ) he p(e-
$i'u5 5e#) i'n5
,
) he 'p) imal pa( ame) e(5 '. ) ') al
.l a$'n'id5 e;) (a)ed *+ &-6 a(e a5 . 'll'45
7 2
mL
'.
2>
e) han'l a5 ) he m'di. ie(
, 3
=/a
, 1
and
3
min '. 5) a)i#
? 12
min '. d+nami# e;) (a#)i'n
.
#
T$e !omparison of SFE (it$ !onventional Ex'
traction
,he #'mpa(i5'n (e5ul) 5 '. &-6 4i) h #'n$en-
)i'nal e;) (a#)i'n unde( ) he 'p) imal #'ndi) i'n5 a(e
li5) ed in , a*le
2.
,he (e5ul )5 5h'4 ) ha) ) he e;) (a#-
)i'n +ield '. &-6 i5 *e) )e( ) han ) ha) '. #'n$en) i'nal
e;) (a#) i'n
.
,hu5 )he p(e5en) &-6 me) h'd i5 u5e.ul
.'( ) he e;) (a#)i'n '. . la$'n'id5 . ('m #p imedium
$ oreanum Na%ai .
Table 2 T$e comparison of total flavonoids extracted by
t$e conventional met$od and by supercritical
carbon dioxide extraction
&le
C'n$en) i'nal
e;) (a#) i'n
&-6
1 1. 1A> 1. 21>
2 1. 2G> 1. 23>
3 A. BA> 1. AB>
% 1. 1G> 1. 1G>
A 1. %> 1. 3%>
A$e(age $al ue5 1. 22> 1. A1>
&) anda(d de$ia) i'n . 2G> . 21>
"ela) i$e 5 ) anda(d de$ia) i'n %. 1> 3. 2>
!onclusions
,he e.. e#) 5 '. m'di. ie(, p(e55u( e, )empe(a)u(e
and e;) (a#) i'n )ime 'n ) he e;)( a#) i'n e. . i#ien#+ '.
) ') al .la$'n'id5 . ('m #p imedium $ oreanum Na%ai
ha$e *een in$e5) iga) ed. ,he &-6 me) h'd i5 #'n5id-
e( ed )' *e $e(+ e. .i#ien) .'( ) he e;) (a#)i'n '. ) ') al
.l a$'n'id5, 4hi#h i5 qui#Fe(, l e55 ) ';i# ) ' ) he en$i-
('nmen) and m'(e e. .i#ien) ) han #'n$en) i'nal me)h-
'd5.
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