Li cm n

Em xin by t lng cm n su sc ti c gio V Th

Ho tn tnh h-ng dn, gip em trong qa trnh chn
v thc hin ti. Em cng xin cm n ban ch nhim
Khoa Ho - Tr-ng i Hc Vinh, cc thy c gio, cc cn
b trong khoa, trong t b mn Ho Phn Tch, Phng th
nghim ho phn tch to iu kin thun li em thc
hin tt ti ny!
Em xin gi ti cc thy c li cm n chn thnh nht.

Sinh Vin: L Th Sen

Lp B2 - K41 - Khoa Ho - i Hc Vinh

Kho lun tt nghip

Li m u
t lu ng-i ta bit -c trong thc n bao gm cc cht dinh
d-ng: protit, gluxit, lipit v mt s mui khong. Nh-ng sau ng-i ta
thy mt s bnh gy ra do thiu mt cht no trong thc n, chnh l
vitamin (theo ting La Tinh: Vita l s sng, vitamin l amin ca s sng).
Vitamin l nhng cht hu c c bn cht ho hc rt khc nhau, m
ch cn mt l-ng nh trong thc n ca ng-i v ng vt bc cao cng
m bo s sinh tr-ng v pht trin bnh th-ng ca c th. Nh-ng vi
mt l-ng rt b nh- vy chng khng phi l nhng cht cung cp nng
l-ng. Ngy nay ng-i ta r vai tr xc tc ca vitamin i vi cc qu
trnh bin i trong c th.
Theo y hc, cc loi vitamin rt cn thit cho hot ng sng ca bt
k c th no, lm xc tc cho qu trnh sinh ho, gn lin vi vic gii
phng nng l-ng, tham gia cc phn ng ca t bo. Vic cung cp khng
y vitamin cho c th s nh h-ng xu khng nhng i vi s lm
vic ca h thn kinh m cn i vi mt lot cc c quan khc bn trong
c th. V th cho nn iu rt quan trng l khu phn n ca con ng-i
phi c gi tr hon chnh khng nhng v ph-ng din calo, v ph-ng
din cht m m con v ph-ng din vitamin na.
i vi vitamin C ni ring cng c vai tr rt quan trng, n c v
tr c bit trong lich s ca hc thuyt v vitamin. Thiu vitamin C c th
s b bnh hoi huyt, nh h-ng tiu cc n s t v ca c th vi cc
bnh nhim trng khc. C th chng ta hon ton khng t tng hp -c
vitamin C m phi ly t nhng ngun thc n ngoi vo. V vy vic phn
tch vitamin C trong thc phm l iu rt cn thit.
Tuy nhin y l hp cht hu c thuc i t-ng kh phn tch, hn
na trong mu phn tch tn ti nhiu cht hu c v v c cc dng khc
nhau, gy cn tr cho qu trnh nh l-ng. Do cn phi la chn

L Th Sen

Lp: 41B2

Kho lun tt nghip

ph-ng php phn tch hu hiu nht.
C th phn tch vitamin C bng cc ph-ng php: sc k giy, cc
ph, trc quang, ha hc... Trong ph-ng php cc ph l mt trong
nhng ph-ng php c tr-ng nht, -c dng nhiu trong thc t nh
l-ng hng lot. Ph-ng php cc ph p ng -c yu cu phn tch hp
cht v c, hu c vi hm l-ng nh, chnh xc. Tuy nhin do iu kin
phng th nghim hin nay ch-a p ng -c nn chng ti khng s dng
-c ph-ng php ny mc d n c nhng -u im r rt.
Khi nh l-ng vitamin C bng ph-ng php trc quang ng-i ta s
dng mt s thuc th v c nh-: axit photphovonframic, mui titan (IV),
axit photphomolipdic... cc ph-ng php ny th-ng gp kh khn khi
phn tch hm l-ng vitamin C trong hoa qu, v dch qu th-ng c mu
v c cha nhiu thnh phn kh khc gy cn tr cho qu trnh nh
l-ng. khc phc mt s nh-c im , trong ti ny chng ti
chn thuc th 2,6 - diclophenolindophenol nh l-ng axit ascorbic.
Vi cch t vn nh- vy nhim v ca ti ny gm:
- Xc nh hm l-ng vitamin C trong n-c cam p nguyn cht ng
hp (nhp t xtrylia) bng ph-ng php chun oxy ho- kh
(ph-ng php phn tch th tch).
- Tm cc iu kin ti -u cho php nh l-ng bng ph-ng php o
quang.
- Th cc iu kin ti -u chn vo vic phn tch mu t to.
- Phn tch hm l-ng vitamin C trong n-c cam p nguyn cht ng
hp v mt s hoa qu c bn trn th tr-ng thnh ph Vinh hin nay.
- Kim tra, nh gi ph-ng php v cc iu kin phn tch vi qu
trnh thc hin v kt qu thu -c, chng ti hy vng rng lun vn s gp
phn b sung, hon thin cc ph-ng php phn tch vitamin C trong nhiu
i t-ng khc nhau nh- n-c p tri cy nguyn cht ng hp v mt s
hoa qu c bn trn th tr-ng hin nay.

L Th Sen

Lp: 41B2

Kho lun tt nghip

phn I
Tng quan ti liu

I - Gii thiu v Vitamin C

1 - L-c s [1].
Bnh Scocbut v cht gy ra bnh ny -c bit t nhng th k
XI - XII. Nh-ng ti khi Holst nghin cu k v c ch tc dng v nguyn
nhn gy bnh, ri Zilva tch -c cht ny t chanh, cam th s hiu bit
v s dng cht ny mi c c s chc chn. Nm 1920 rumlnond t tn
cht ny l vitamin C. Vi nm sau, Her - Bert. Hirst v Karrer tch -c
vitamin C t th-ng thn. Nm 1933 Funk ri Reichstein, Haworth tng
hp -c vitamin C. Ngy nay ng-i ta bit vitamin C c trong t nhin
d-i dng: kh (axit L. Ascorbic) v dng oxy ho (axit dehydroascorbic),
dng sau ny t hn. C hai dng u tan trong n-c, d b phn hu khi
tip xc vi cht oxy ha hoc baz. So vi cc vitamin khc th vitamin C
l cht duy nht khng c dng phc hp vi cc Nucleotit hoc
Coenzym. Hin nay ng-i ta iu ch vitamin C bng tng hp r hn
nhiu so vi ph-ng php iu ch t thc vt.
2 - Vai tr ca Vitamin C trong i sng con ng-i [1, 2, 3].
Vitamin C c v tr c bit trong lch s ca hc thuyt v vitamin.
Vitamin C tham gia vo cc qu trnh oxy ho - kh khc nhau c th. N
xc tc cho s chuyn ha nhiu hp cht thm thnh cc dng Phenol t-ng
ng V d: Qu trnh hydroxyl ho triptophan thnh hydroxytriptophan, hoc
phenylalanin chuyn thnh tirozin. Phn ng chuyn amin ho gia tirozin v
axit - xetoglutaric to nn sn phm l axit para oxyphenylpiruvic cng
thc hin vi s tham gia ca vitamin C. Ngoi ra vitamin C cn tham gia
iu ho s to ADN t ARN hoc chuyn procolagen thnh colagen. Nh
qu trnh hyroxyl ho prolin to nn cht oxyprolin cn thit cho s tng

L Th Sen

Lp: 41B2

Kho lun tt nghip

hp colagen. Chnh v vy n c tc dng lm cho vt th-ng chng lin so.
vitamin C cn lin quan vi s hnh thnh cc hormon ca tuyn gip trng
v tuyn trn thn. N rt cn thit cho c th tng sc khng v chng
li cc hin t-ng chong hoc ng c bi cc ho cht cng nh- cc c t
ca vi trng. Ng-i ta cng chng minh rng vitamin C c lin quan ti s
trao i gluxit c th. V khi b bnh hoi huyt trao i gluxit c tim b
ri lon s phn gii glicogen v glucoza tng ln mnh, ng thi tng tch
lu axit lactic. Hin t-ng ny s bin i nhanh chng nu thm vitamin C
vo cc cht dinh d-ng.
Khi thiu vitamin C trong thc phm th ng-i ta s b bnh hoi
huyt. V bnh hoi huyt pht trin t t nn vic chun on trng thi
thiu vitamin C rt kh khn. Bnh thiu vitamin C -c c tr-ng bi s
gim tnh chu ng ca c th i vi cc bnh nhim trng khc nhau.
Bnh nhn rt chng mt hay au u v lun lun trng thi mng m,
n km ngon. Sau nhp p ca tim b ri lon, thnh cc mao qun tr
nn mng manh v d b t, do c th xy ra cc tr-ng hp chy mu
t pht. Khi b bnh thiu vitamin C, ngay c trong nhng tr-ng hp c
nhng tc ng rt nh v c hc cng c th gy ra hin t-ng chy mu
cc c quan bn trong. Trong bnh thiu vitamin C cng km theo nhng
thay i rt c tr-ng v x-ng v rng: Rng t phn hy v rng rt
nhanh. Bnh thiu vitamin C cn -c gi l bnh scobut.
Nhng ri lon trong vic tng hp cht colagen l c s ca nhng
hin t-ng trn. iu cng gy nn nhng thay i bnh l ca cc
thnh mao qun v ca cc m chng . Hng ngay c th cn mt l-ng
vitamin C nhiu hn cc loi vitamin khc. Nhu cu vitamin C thay i theo
tui, kh hu v lao ng. Ng-i ln mi ngy cn khong 50 - 100mg. i
vi ph n c thai hoc tr con mi ngy cn 100 - 200mg.
Tm li vitamin C rt cn thit i vi c th con ng-i. Tuy nhin trong
c th chng ta hon ton khng th t tng hp -c vitamin C, m phi ly t
ngun thc n bn ngoi. Vy vitamin C trong thin nhin tn ti u?

L Th Sen

Lp: 41B2

Kho lun tt nghip

3 - Cc ngun Vitamin C trong thin nhin [1, 2, 3, 4]
Vitamin C tn ti trong thin nhin d-i 3 dng ph bin l axit
ascorbic (dng kh); axit dehydroascorbic (dng oxy ho) v ascorbigen
(dng kt hp).
Vitamin C c nhiu trong cc loi rau qu nh-: Cam, chanh, du, t,
rau ci, c chua... . Cn trong cc loi ht ng cc hoc trong trng, tht hu
nh- khng c vitamin C.
Hm l-ng vitamin C bin i nhiu ph thuc vo loi, v tr trng
trt v cc yu t nh- chiu sng, kh hu... Bnh th-ng l-ng vitamin C
gim dn t pha v ngoi vo bn trong rut ca qu vitamin C xut hin
hu ht cc b phn ca cy mm gp nhiu h: n, thng, ci, t gp
h: cc, trch, t.
.4 - Cng thc ho hc:
4.1 - Axit L - Ascorbic
- Cng thc phn t:
- Cng thc cu to.

C 6 H8 O6
O
C
C

OH
O

C
H

HO

OH

CH2OH
Tn thng th-ng: Axit ascorbic; vitamin C; cevitamin axit;
hexuromic axit
- Tn ho hc: L () Threo - 2, 3, 4, 5, 6 Pentahydroxyl - 2 hexanoic - axit - 4 - Lactone.

L Th Sen

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Kho lun tt nghip

4.2 - Axit L - dehydroascorbic:
- Axit L - dehydroascorbic cn gi l vitamin C2.
- Cng thc phn t: C6H606.
- Cng thc cu to:
O
C
C

O
O

C
H

HO

CH2OH
- y l dng oxy ho.
5 - Ph-ng php iu ch [5]
Ngy nay ng-i ta iu ch vitamin C bng con -ng tng hp ho
hc, ph-ng php ny r hn nhiu so vi ph-ng php iu ch vitamin C
t thc vt. Nguyn liu l D - glucose. Hydrogen ho D - glucose (1) vi
xc tc Cu - Cr -c D - sorbitol (2). Trong dung dch n-c, D - sorbitol
d-i tc dng ca vi khun Acetobacter suboxydans to ra L - sorbose (3).
Ng-ng t (3) vi axeton vi xc tc axit sunfuric to diaxeton sorbose (4).
Oxy ho (4) bng kalipermanganat to (5); thu phn (5) to axit 2 ceto L gulonic (6). Este ho (6) bng methanol, sau cho tc dng vi natri
methoxyd to (7). Thu phn (7) trong dung dch axit hydrocloric trong
n-c loi nhm methyl v natri, lacton ho to axit ascorbic (8):
CHO
CH2OH
CH2OH
CHOH

CHOH

C=O

H2
HOCH

A. Suboxydans
HOCH

HOCH

CHOH

CHOH

CHOH

CHOH

CH2OH

CH2OH

(1)

L Th Sen

CHOH
HOCH
CH2OH

(2)

(3)
6

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Kho lun tt nghip

H3 C

CH3

H3 C

CH3

C
H

O
H2 O

KMnO4
H
H

H2)HH2O

H2C

H2 C

COOH

CH2OH
O

H3 C

CH3

H3C

(4)

CH3
(5)

O
COOH
+ H2 O

COOCH 3
CH3OH + HCl

C=O

C
HCl

HOC

OH

Sau CH3ONa
HO - CH
CH OH
HO - CH
CH2OH
(6)

O
NaOC
CHOH
HOCH
CH2OH
(7)

C
H

HO

OH

CH2OH
(8)

6 - Tnh cht ca vitamin C [3, 4].

6.1 - L tnh.
Axit L - ascorbic(C6H8O6) kt tinh hnh mnh, khng mu, v chua.
Nng chy nhit 190C n 192C, tan hon ton trong H2O (1g trong
3ml n-c, trong 50ml cn). Khng tan trong benzen; clorofoc; ete v m.
trng thi kh, kh vng bn trong khng kh, dng dung dch th nhanh

L Th Sen

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chng b phn hu khi c khng kh, c bit trong mi tr-ng kim. y
l cht c hot tnh quang hc []D20 = + 23 (trong H2O).
.6.2 - Ho tnh.
V ho tnh, l ho tnh ca chc lacton, ca cc nhm hydroxyl.
Song quan trng nht l ho tnh ca nhm endiol. Nhm ny gy ra tnh
axit v tnh kh ca axit ascorbic.
.6.2.1 - Tnh axit.
Do hiu ng lin hp vi nhm cacbonyl nn nguyn t hydro ca
nhm hydroxyl v tr s 3 tr nn rt linh ng lm cho vitamin C c tnh
axit mnh. V vy vitamin C c tn l axit ascorbic.

O:

OH

Do c tnh axit mnh nn axit ascorbic d tan trong cc dung dch

kim cng nh- cacbonat kim loi kim. ng dng tnh cht ny, ng-i ta iu
ch mui ascorbat natri pha dung dch tim c nng cao v trung tnh.
Tc dng vi mui kim loi cho mui mi. Nhiu d-c in dng
thuc th l st (II) sunfat hoc st (III) clorua nh tnh ascorbic
Ph-ng trnh phn ng:
CH2OH

CH2OH

HOCH

HOCH
O

O
O

HO

L Th Sen

+ NaHCO3
Sau + FeSO4

OH

O Fe

O
8

OH
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Kho lun tt nghip

6.2.2 - Tnh kh l c tnh c bn ca Axit ascorbic.
Axit ascorbic cng nh- tt c cc endiol c tnh cht kh mnh v b
oxy ho trong mt phn ng thun nghch thnh axit dehydroascorbic. C
th dng cc thuc th sau y oxi ho: oxy, dung dch it; 2,6 diclophenolindophenol; xanh metylen; kaliferixianua... Tc ca s oxy
ho trong cc mi tr-ng ph thuc vo pH, pH cc i l 5,0; vi phn ng
c baz, pH cc i l 11,5. Trong mi tr-ng kim phn ng oxi ho xy
ra nhanh hn trong mi tr-ng axit.
Cc tc nhn xc tc s oxy ho l: nh sng, nhit , cht kim, cc
enzym v vt ng hay st.
S oxy ho kh thun nghch axit ascorbic thnh axit dehydroascorbic.
O

OH

[O]

C=O

O
C
H

HO

OH

O
[H]

C=O

HO

CH2OH

CH2OH

Axit ascorbic

Axit dehidroascorbic

Tnh cht ny v cng quan trng i vi tc dng sinh hc ca axit

ny. Trong c th, axit ascorbic c th cho hai nguyn t hydro (l cht
cho) to thnh axit dehydroascobic; dng ny, n c th nhn hai
nguyn t hydro (l cht nhn) tr li axit ascorbic. Nh- vy axit
ascorbic tham gia vn chuyn hydro tc l tham gia vo cc h enzym xc
tc cc qu trnh oxy ho kh xy ra trong c th .

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Axit dehydroascorbic vng bn trong mi tr-ng axit PH d-i 4,0. D b
phn hu trong mi tr-ng trung tnh v kim thnh axit 2,3 dixetogulonic khng c tc dng sinh hc.
O
C

COOH

C=O

C=O

+ H2O

C=O
H

HO

C=O

OH

HO

CH2OH
Axit dehydroascorbic

CH 2OH
Axit 2,3 - dicetogulonic

II - Cc ph-ng php xc nh Vitamin C:

1 - Nhn xt chung:
Vic phn tch vitamin C trong thc phm l iu rt cn thit phc v
cho vic nh gi gi tr thc phm i vi i sng con ng-i. Cc ph-ng
php xc nh vitamin C c th chia lm hai nhm:
+ Ph-ng php ho l
+ Ph-ng php ho hc.
Trong nhng ph-ng php ho l th ph-ng php cc ph c mt
ngha thc t. Trong nhng nm gn y ph-ng php ny rt -c pht
trin v -c dng nghin cu vitamin C v mt l thuyt. Ph-ng
php sc k giy vi nhiu kin thc mi qu gi -c s dng
nghin cu axit ascorbic dng kt hp. Trong cc ph-ng php o mu,
nhiu ph-ng php c ngha nht u da trn phn ng ca axit ascorbic
dng oxy ho vi 2,6 - dinitrophenylhydrazin. Sau y l tm tt v cc
ph-ng php ho hc v vt l c th s dng xc nh vitamin C

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2 - nh l-ng axit ascorbic bng ph-ng php vt l:
2.1 - Ph-ng php quang ph [4]:
Axit ascorbic c ph hp th c tr-ng vng t ngoi cc i c
b-c sng 265 nm; log = 3,98 (trong mi tr-ng n-c). Khi ghi ph cn
phi lm nhanh cht kh khi b oxy ho. Mt s tc gi ngh dng
mi tr-ng cha ion xianua v nh l-ng bng quang ph dung dch
axit ascorbic. V tr ca di hp th ch yu ph thuc vo tnh cht ca
dung mi, th d trong n-c cc i 265nm, trong etanol 245nm. V tr
cc i ph thuc vo pH v khi pH gim i th s dch v nhng b-c sng
ngn hn. S thay i ph hp th ca axit ascorbic i vi pH ph thuc
vo trng thi cn bng trong dung dch gia dng endiol v dng xeton. V
mt quang ph, axit ascorbic c tnh cht t-ng t nh- cc endiol khc, cc
axit kh, cc cht kh... ph hp th t ngoi ca dinitrophenylhydrazon,
axit dehydroascorbic v cc sn phm endiol oxy ho khc -c dng
nh tnh cc cht ny. Ph hp th ca axit ascorbic c th dng nh- mt
hng s vt l c tr-ng v th nh tnh khi nh gi cc cht dng lm
mu chun hoc dng nh l-ng axit ascorbic trong dung dch tim.
Zprochazka o ph hp th ascorbigen trong vng t ngoi khi
nghin cu cu to ca cht ny. So snh cc ph ca ascorbigen, indol v
axit ascorbic th ph ca ascorbigen khng ging ph axit ascorbic t c
th suy ra l axit ascorbic khng kt hp dng endiol. Ph t ngoi ca
ascorbigen thc t trng vi ph ca indol, t c th suy ra l trong phn
t ascorbigen c vng hoc c cc nhm c cu trc t-ng t.
2.2 - Ph-ng php cc ph [4].
Cc ph nghim l mt trong nhng ph-ng php c tr-ng nht
-c dng nhiu trong thc t nh l-ng hng lot. Axit ascorbic c
tnh cht cc ph ging nh- cc endiol khc v cho sng ant i in t
khng thun nghch. Tuy rng sng cc ph ca axit dehydroascorbic c
tnh kh, nh-ng n nhiu ln thp hn sng ant t-ng ng ca axit

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ascorbic. C ch kh ny thc ra ch-a r rng. Tnh cht ca axit ascorbic
khi nh l-ng bng ph-ng php oxy ho v s thay i th nng chun
ca n i vi pH cho php gi thit rng dng endiol c kh nng trao i
thun nghch i in t vi in cc. Kwiesner theo di qu trnh oxy
ho thun nghch in cc khi nghin cu axit dehydroxyfumaric bng
cc ph giao ng. Tc gi cho rng cht ny c cha nhm endiol, u
tin cho sn phm oxy ha thun nghch khng bn vng, sau sn phm
ny hydrat ho thnh sn phm khng thun nghch khng bn vng.
S chng minh t-ng t i vi ascorbic khng c kt qu c th do
i sng ngn ngi ca sn phm thun nghch.
S oxy ho axit ascorbic bng ph-ng php cc ph trn in cc
thu ngn nh git khc bit vi ph-ng php in th ch th nng oxy
ho cc ph khong 200mv d-ng hn gi tr thu -c vi ph-ng php
o in th. Theo J.Heyrovsky v Z. Vavrina th trong s di chuyn ny c
dng sng t-ng t nh- trong qu trnh thun ngch; bng cch ng dng
ph-ng php tnh tc phn ng (K) sau y, th dng oxy ho in hot
s chuyn thnh khng in hot (V):
K

As 2e + 2H+ + DA*

DA

Trong :
As = axit ascorbic
DA* = Sn phm oxy ha thun ngch u tin.
DA = Sn phm oxy ha th hai sau khi hydrat ha.
K = Hng s tc hydrat ho
Gi thit cho rng s bin i axit ascorbic thnh axit
dehydroascorbic khng hydrat ha l thun ngch v s hydrat ho ca sn
phm oxy ho th nht trn in cc b chm do qu trnh ring ca in
cc, -c gii thch bng s t-ng quan gia s di chuyn th bn sng vi
thi gian nh git, iu ny ph hp vi cc kt qu thc nghim. Nm

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1933 E. Kodicek v K. Wenig s dng ph-ng php cc ph thm d
nh l-ng axit ascorbic t do. H nghin cu trong mi tr-ng photphat
c m pH = 7,0 v nh l-ng trong dch qu chanh, qu cam bng cch
trn n gin dch vi m theo t l 1 : 1. Ghi -ng cong sau khi
loi oxy ho tan bng cch sc kh tr. Ph-ng php v dung dch m
trn -c I.okada dng nh l-ng axit ascorbic trong dch hoa qu
hp. K.Schwars ngh thay m photphat bng m axetat pH = 4,7
vi l do l axit ascorbic trong mi tr-ng axit th vng bn hn trong mi
tr-ng trung tnh, v tin hnh nh l-ng bng cch trn dch vi
dung dch m theo t l 1 : 5. nh l-ng axit ascorbic trong d-c
phm, O. Mata ngh dng m phtalat c axit oxalic pH khong 4,0. Vi
cch lm nh- trn th vic xc nh khng b tr ngi khi c mt aspirin;
histidin; vitamin K3; pyridoxin...
P. Zuman l-u ti nhiu chi tit trong vic nh l-ng bng cc
ph axit ascorbic bn cnh cc cht sunfhydryl th-ng c trong cc m
thc vt d-i dng glutathion; cystein; ergothionein... v cho sng ant d
nhm ln vi sng ca axit ascorbic. Cc cht ny c nh h-ng khng tt
n vic nh l-ng axit ascorbic bng ph-ng php oxyho, bi v cng b
oxyho vi axit ascorbic v lm tng l-ng thuc th oxyho. P. Zuman
tm thy cc iu kin thch hp c th tin hnh nh l-ng ng thi
axit ascorbic v cc cht Sunfhydrin. Cch lm n gin v thc t tc gi
ngh dng m axetat pH = 4,7 lm mi tr-ng thch hp nht.
Mt s tc gi ch n vic nh l-ng vitamin C trong nhng
nguyn liu rt giu axit ascorbic. H l-u n vitamin C c km theo
cc cht khc c tnh kh mnh, gy cn tr cho nh l-ng. M. Pyke, R.
Melville v H. Sarson ri C. Daglish v F. Wokes xc minh bng o
quang ph l cht kh ny l este ca hydrojuglon m trng c v nh- axit
ascorbic.

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O

OH

+ 2H+ + 2e

OR

OR

OH

phn bit c hai cht c th dng phn ng vi focmandehit, c

bn l hydrojuglon khng tc dng vi focmandehit trong mi tr-ng axit
mnh. Ng-i ta cng nghin cu ph-ng php da trn tc oxyho
khc nhau ca hai cht bng oxy cc pH khc nhau.
2.3 - Ph-ng php sc k [4].
Ph-ng php sc k giy mang li nhiu kt qu qu cho vic
nh l-ng axit ascorbic khi c mt cc cht cn tr. N cho php pht hin
nhanh chng cc thnh phn cn tr khc nhau v nh tnh -c chng. Do
c th la chn t-ng i d cnh nh l-ng thun li, s ci tin thch
hp L. M. Mapson v S. Partridge nghin cu ph-ng php ny c kt
qu. H dng

h dung mi: phenol bo

ho n-c, colidin

bo

ho n-c v h: n- butanol - axit axetic - n-c (4 : 1 : 5) pht hin bng

dung dch 2,6 - diclophenolindophenol. Z. Prochazka dng giy whatman
s 1, h dung mi metanol - n-c v pht hin bng dung dch bc nitrat
amoniacat hoc dung dch h tinh bt c iodua. V.Sanda kho st nhiu
cch khc nhau xc nh hm l-ng axit ascorbic trong cht liu thc vt
bng ph-ng php sc k giy v nghin cu ci tin tt vic nh l-ng
trong thc phm. Cc h dung mi -c dng l: metanol trong n-c 50%,
h butanol - axit axetic - n-c (50ml n-c, 40ml n- butanol v 14,5ml axit
axetic bng cho n khi mt hn gii hn ngn cch ca hai pha). Tip
h dung mi do Mapson ngh c kt qu tt: n - butanol - axit axetic n-c (4 : 1 : 5) pht hin bng dung dch 2,6 - diclophenolindophenol.

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Chy sc k theo h-ng t d-i ln trn trong kh quyn dioxit
cacbon nh l-ng bng ph-ng php cho thm cht chun. Kt qu thu
-c trong ph-ng php sc k -c kim tra li bng ph-ng php
oxyho.
H.V. Euler v cng s dng ph-ng php sc k giy theo di
s bn vng ca axit ascorbic trong nhng mi tr-ng khc nhau. Ph-ng
php sc k giy -c dng nh l-ng axit ascorbic trong n-c tiu
v cc t chc, theo di s to ra cc cht kh trong -ng... Ph-ng
php s dng rt tt nh l-ng axit ascorbic kt hp trong cy xanh.
Z.Prochazka thm d 26 h dung mi khc nhau nh l-ng
ascorbigen trong rau xanh v ngh dng cc h: etylaxetat - n-c; butyl
axetat - n-c, n - butanol - n-c. Hn hp gm focmaldehyt v axit
clohydric ngh lm thuc th pht hin ascorbigen, n cho vi
ascorbigen mt vt mu da cam c mu hunh quang sm trong nh sng t
ngoi. Sau khi thu phn ascorbigen dng thuc th h tinh bt c iodua
pht hin axit ascorbic t do. Sc k trn giy whatman theo chiu t d-i
ln.
F. Samraj chn mt cch sc k giy khc nh l-ng
ascorbigen, lm trong kh quyn hydro v dng hn hp gm 50ml n-c,
40ml n - butanol lm h dung mi v thm 100ml axit axetic bng cho ti
khi hn hp nhuyn u. Pht hin bng mt dung dch bc nitrat 1% trong
axit axetic 2%. Tc gi ng dng vo qu kim anh, bp ci v t ngt.
Trong tt c cc mu th thy ascorbigen bn cnh axit ascorbic. C th
phn bit -c chng khng ch theo gi tri Rf, m cn theo hnh dng vt
sau khi pht hin. Trong thc t theo di hm l-ng ca hai cht trong
mu th tr-c v sau khi thu phn trong mi tr-ng axit axetic 10%. Cc
kt qu ny -c cc tc gi kim tra li bng ph-ng php oxy ho v
cc ph. Vic xc nh l-ng axit ascorbic trong cht liu sinh vt -c
Yu - Tuan - Chen v cc cng s l-u ; bng ph-ng php m t, h c
th tch axit L - ascorbic ra khi axit D - araboascorbic. Sau khi trin khai

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sc k , em ct theo tng vt v chit tng vt ri xc nh gi tr bng
ph-ng php so mu sau khi cho tc dng vi diclophenolindophenol. Vic
xc nh axit dehydroascorbic bn cnh axit ascorbic bng ph-ng php sc
k giy -c H. Smidt v H. Staudinger nghin cu.
tch ascorbigen t dch m c thu -c t bp ci, Z. Prochazka
dng sc k phn chia trn ct xenluloza v thu -c dch m c
ascorbigen rt tinh khit c cha 31% axit ascorbic. trin khai sc k
dng h ete - n-c.
3 - nh l-ng axit ascorbic bng ph-ng php ho hc [4].
3.1 - nh l-ng bng ph-ng php oxy ho [4].
Cng nh- trong cc ph nghim c s nh-ng in t cho in cc,
y sau khi thm thuc th oxy ho vo dung dch axit ascorbic th s
oxy ho c th xy ra. Ng-i ta dng iot; kaliferixyanua; 2,6 diclophenolindophenol, xanh metylen... lm thuc th oxy ho. C th theo
di s chun oxy ho axit ascorbic bng mt, o quang, o in th v
o cc ph. Do tnh khng bn vng ca axit ascorbic ngoi kh tri nn
cn c bit l-u khi pha ch dung dch v chit t cht liu thin nhin.
Vic nh l-ng axit ascorbic bng ph-ng php oxy ho thng th-ng tin
hnh bng cch nh t buret dung dch thuc th vo dch chit axit
ascorbic trong mi tr-ng thch hp axit metaphotphoric hoc cc axit thch
hp khc. Trong tr-ng hp n gin nht, ng-i ta quan st s mt mu
bng mt. Trong php chun bng o in l-ng, ng-i ta dng bch
kim hoc nhng in cc khc ch to mt cch c bit lm in cc ch
th. Php o in l-ng t-ng i t dng nh l-ng axit ascorbic trong
cht liu thin nhin v n t c tr-ng v th chm n nh. Php o quang
rt hay -c dng quan st s oxy ho axit ascorbic bng 2,6 diclophenolindophenol, trong s mt mu dn -c theo di bng tt
o b-c sng 520nm. Ng-i ta nghin cu ci tin ph-ng php ny
bng cch theo di tc mt mu, t suy ra -c tnh cht ca cc tp
cht kh c mt. Mt trong nhng ci tin ny do F. Follkmann nghin cu

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l vic loi nh h-ng ca tp cht kh km theo.
Vic ci tin da trn nguyn tc chit 2,6 - diclophenolindophenol
ch-a b kh trong mi tr-ng phn ng axit bng xylen. E. Stotz da trn
ph-ng php c bn ny nghin cu nh l-ng axit ascorbic trong mu.
Theo cch lm trn ng-i ta c th ngn tr s xc tip ca dng oxy ho
qu tha ca cht mu vi cc cht tc dng chm, tr axit ascorbic. Trong
phn trnh by d-i y l nhng ph-ng php hay dng nht nh l-ng
axit ascorbic da trn ph-ng php oxy ho v bng nhng th d s minh
ho nhng ci tin khc nhau tin hnh trn nhng cht liu nht nh.
3.1.1 - Ph-ng php chun bng 2,6 - diclophenolindophenol quan
st bng mt.
Khi chun oxy ho axit ascorbic bng 2,6 - diclophenolindophenol
trong mi tr-ng axit, axit ascorbic b oxy ho thnh axit dehydroascorbic
v cht mu b kh thun nghch thnh dng khng mu. S kt thc nh
l-ng -c bo hiu bng s xut hin mu hng, chng t s c mt ca
mt l-ng c tha cht mu khng b kh.
Ph-ng php c th dng nh l-ng trc tip axit ascorbic trong
cc ch phm d-c v vi ci tin thch hp c th nh l-ng trong thc
phm. Vic ci tin ch yu nhm loi tr nh h-ng ca nhng cht cn
tr nh- cc cht kh, cc cht sunfhydryl. Bng cch cho cc cht
sunfhydryl ng-ng t vi focmandehit iu kin thch hp.
Nh- vy xc nh gi tr dch chit cn phi tin hnh 3 ln nh
l-ng iu kin khc nhau theo s sau:
1 - Chun khng c focmaldehit xc nh axit ascorbic, cht
kh v cc cht sunfhydryl ni chung (A)
2 - Chun c focmaldehit trong mi tr-ng pH = 0,6 xc nh
axit ascorbic v cc cht kh (B) iu kin ny nh h-ng ca cc cht
sunfhydryl -c loi bng focmaldehit.
3 - Chun trong mi tr-ng m pH = 3,5 vi s c mt ca
focmaldehit th ch xc nh -c cc cht kh (C).

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3.1.2 - nh l-ng axit ascorbic trong cht liu thin nhin bng ph-ng
php o quang [4].
Ph-ng php o quang da trn s theo di qu trnh lm mt mu
ca 2,6 - diclophenolindophenol bng dung dch axit ascorbic chun v
bng dch chit. S mt mu do axit ascorbic thc t xy ra tc thi, cn s
mt mu do cc cht tp c tnh kh phn nhiu chm hn. Do c th
tin hnh iu chnh bng cch theo di thi gian qu trnh xc nh hm
l-ng thc ca axit ascorbic.
Ph-ng php ny thch hp cho nhng tr-ng hp dung dch trong
v hm l-ng axit ascorbic thp. Khng th ng dng cho dung dch c
mu hoc dch chit c nhiu. Trong tr-ng hp ny nn dng ph-ng
php indophenol ci tin dng xylen.
3.1.3 - nh l-ng bng o quang axit ascorbic theo ph-ng php chit
indophenol - xylen [4].
Sau khi x l cht liu cn th, tc gi xem cht liu c cha cht
mu chit -c bng xylen khng. Th nghim bng cch trn 5ml dch
chit axit, 5ml m v chit vi 10ml xylen. Nu tt ca dung dch ln
hn 0,01 th cn phi dng gi tr ca mu trng sau khi thm hydroquinol.
Tin hnh kh cht mu bng axit ascorbic, chit l-ng tha ca
cht mu bng xylen. Sau em o mu, trn c s chng ta nh
l-ng -c axit ascorbic.
3.1.4 - Chun axit ascorbic bng ph-ng php o in [4].
Chun bng ph-ng php o in c -u im r rt khi nh l-ng
axit ascorbic trong dung dch c hay c mu m, y khng th dng
ph-ng php nhn bng mt hoc so mu -c. E. Becker v J. Gleria dng
bch kim nhn lm in cc ch th, in cc calomen lm in cc so snh.
H tin hnh nh l-ng trong dung dch nhm sunfat M. Kirk v D.
Tessler dng in cc l-ng kim (bch kim v antimon). Vic chun tin
hnh trong mi tr-ng axit axetic 8% L. Harris v L. Mapson kho st t

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m vic chun in th axit ascorbic v ngh dng nhng in cc
bch kim c bao thu ngn. Khi ng dng ph-ng php ny vo vic inh
l-ng cc cht thin nhin c mt ca nhng cht cn tr lm cho th chm
n nh v im cui khng r rng, do chnh xc b gim i. Ng-i
ta c th dng ph-ng php in th nh l-ng axit ascorbic trong ch
phm d-c.
Chun o in axit ascorbic vi 2,6 - diclophenolindophenol theo
L. Mapson, L. Harris v Y. Wang: tc gi dng in cc bch kim bc thu
ngn lm catt v bch kim bng lm ant. Chun trong mi tr-ng axit
metaphotphoric 5% bng dung dch 2,6 - diclophenolindophenol.
3.2 - nh l-ng axit ascorbic bng ph-ng php o quang [4].
Axit ascorbic cho mt lot phn ng mu c th dng nghin cu
nh l-ng cht bng ph-ng php so mu. M.Bachstez v G.Gavallini
m t axit ascorbic cho mu nu khi c mt uranylaxetat trong mi
tr-ng kim nh. N. Bezssonoff dng ph-ng php so mu nh l-ng
axit ascorbic nh mu xanh l m axit ny to nn vi axit
photphovonframic trong nhng iu kin thc nghim nht nh. E. Sehulec
v J. Floderer m t mt phn ng mu rt nhy. Vic nh l-ng
tocoferol theo Emmre v engel cng da trn mt phn ng t-ng t. Vic
nh l-ng da trn cch o mu thm xut hin khi c tc dng ca ,
, - dipyridyl trn ion st ho tr II, ion ny sinh ra do axit ascorbic kh st
III clorua. Xt v bn cht ca ph-ng php th r rng l phn ng rt
khng c tr-ng. J. H. Roe v C. H. Kuther gii thiu nhiu ln ph-ng
php so mu, trong axit ascorbic b oxy ho bi brom hoc iot thnh axit
dehydroascorbic, cht ny tc dng vi dinitrophenylhydrazin thnh osazon
c mu t-ng ng. Nm 1953 M.Schmall, CH. Pifer v E. Wollisch
nghin cu mt ph-ng php mi da trn cch o mu xanh l sinh ra khi
cho tc dng diazo - 4 - metoxy - 2 - nitroanilin trn axit ascorbic. Ph-ng
php -c coi l kh c tr-ng nn n -c dng nh l-ng axit

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ascorbic trong polyvitamin v trong qu v s -c trnh by y d-i.
3.2.1 - nh l-ng axit ascorbic,dehydroascorbic v dixeto - gulonic bng
ph-ng php vi 2,4 - dinitrophenyldrazin.
Theo J. H. Roe v M.B.Mills th ph-ng php ny c th dng
nh l-ng cc axit ascorbic, dehydroascorbic, dixetogulonic trong cht liu
thin nhin, n sinh ra bng cch thu phn axit dehydroascorbic. Axit
dixetogulonic khng c tc dng sinh l v ng-i ta xc nh rng hm
l-ng ca n tng ln trong qu hay rau xanh l-u kho. Ph-ng php da
trn vic nh l-ng bng cch o mu sinh ra do tc dng ca 2,4 dinitrophenyldrazin trn axit dehydroascorbic v dixetogulonic. Bng cch
b tr thc nghim thch hp, ph-ng php ny c th nh l-ng -c c
ba thnh phn bn cnh nhau. Kt qu ph thuc vo tc th nghim, bi
v axit dehydroascorbic d chuyn thnh dixetogulonic. Thioure -c cho
thm vo vi mc ch lm n nh dng kh ca axit ascorbic.
3.2.2 - nh l-ng axit ascorbic bng so mu theo M.Schmall, Chpifer v
E.G. Wollisch.
Da vo o sn phm mu ca phn ng gia axit ascorbic vi 4 metoxy - 2 - nitroanilin diazo ho. Trong mi tr-ng kim, cht ny ho
tan v cho mt mu xanh thm, ph-ng php ny kh c hiu v c th
cho php nh l-ng axit ascorbic bn cnh cc sn phm oxy ho, ngha l
bn cnh axit dehydroascorbic v 2,3 - dixetogulonic. C th dng ph-ng
php ny nh l-ng trong thc phm v hoa qu.
III. c im ca thuc th hu c 2,6 - diclophenolindophenol
2,6 - diclophenolindophenolat natri C12H6O2NCl2Na.
Tnh cht: l bt mu xanh l cy thm, tan trong n-c v r-u etylic.
Dung dch etylic ca thuc th c mu xanh, khi axit ho tr thnh , pH = 5,7.
Phn ng: Khi thm thuc th vo dung dch KI trong mi tr-ng axit
th c it thot ra. Vi tc dng ca cc cht kh mnh nh- axit ascorbic th
mu xanh ca thuc th chuyn thnh mu vng nht gn nh- khng mu.

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ng dng: xc nh axit ascorbic, dng lm ch th oxy ho kh.
E0 = + 0,217 V pH = 7 (dng oxy ho c mu xanh)
E0 = + 0,67 V pH = 0 (dng kh khng mu, dng oxy ho mu ).
IV - Kt lun
Qua nghin cu cc tnh cht ca vitamin, qua phn tng quan v cc
ph-ng php phn tch v cn c vo iu kin thc t ca phng th
nghim chng ti chn thuc th 2,6 diclophenolindophenol nh
l-ng axit ascorbic theo hai ph-ng php:
+ nh l-ng axit ascorbic bng ph-ng php chun oxy ho-kh
(ph-ng php phn tch th tch).
+ nh l-ng axit ascorbic bng ph-ng php o quang.

Nguyn tc chung ca hai ph-ng php:

Ph-ng php da trn nguyn tc l axit ascorbic c kh nng oxy ho kh

thun nghch nh trong phn t ca n c cha nhm endiol.
C

OH

OH

V vy axit ascorbic kh cht ch th c tr-ng mu xanh dng nh

l-ng n l 2,6 - diclophenolindophenol trong mi tr-ng axit n dng
khng mu. Trong khi axit ascorbic li b oxy ho thnh axit
dehydroascorbic.
2,6 - diclophenolindophenol l hp cht c kh nng i mu kp mt
mt khi PH ca mi tr-ng thay i t kim qua axit th mu ca cht ch
th chuyn t xanh sang hng. Mt khc dng oxy ho ca 2,6 diclophenolindophenol c mu, cn dng kh khng mu. Chnh v vy s
c mt ca mt l-ng tha cht mu khng b kh trong mi tr-ng axit s
lm cho dung dch c mu hng.

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S phn ng gia axit ascorbic v 2,6 - diclophenolindophenol
O
C
Cl
C

OH
O

=N

OH

OH
Cl

HO

Dng oxy ho ca 2,6 diclophenolindophenol

c mu xanh

CH2OH
Axit ascorbic

O
C
Cl
C

O
O

HO

NH

OH

O
Cl

HO

Dng kh khng mu

CH2OH
Axit dehydroascorbic
V - X l thng k cc s liu thc nghim.
1 - X l kt qu phn tch:
Trong mt php o c hn (n << ) ta nhn -c cc gi tr : X1, X2,
X3... Xn. Sau khi loi tr sai s th bng chun ixan (Q) ta tnh -c gi

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tr trung bnh X nh- sau:
- Chn trong s cc gi tr thc nghim c mt gi tr no v
t bng C.
-

Tm cc lch ca mi php o so vi gi tr C.
xi =Xi - C

- Tnh gi tr trung bnh theo cng thc:

X=C+

1
n

xi

Ph-ng sai:
S2 =

1
k

(Xi - x )2

k=n-1
lch chun ca gi tr trung bnh:
S2
Sx =
n

Cc kt qu phn tch -c phi x l nh gi chnh xc ca

php xc nh:
= tP,k. S x

tP,k : Hm l-ng Studenst vi xc xut p, bc t do k (tra bng)

Vy khong xc nh ca kt qu l:
X - X +

Sai s t-ng i ca php o.

q% =

.100
X

2 - So snh kt qu phn tch vi mu bit hm l-ng

kim tra mt php phn tch mi c dng -c hay khng th
chng ta tin hnh lm th nghim v nhn -c gi tr X . Sau em so
snh X vi gi tr chun a theo nm b-c sau:
b1: Gi thit rng X a l do nguyn nhn ngu nhin vi xc xut
p = 0,95

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b2: Ln l-t tnh cc tham s X , S x
b3: Tnh tTN theo cng thc:
tTN =

X a
Sx

b4: Tm tlt = tP,k (tra bng)

b5: So snh tlt v tTN:
Nu tP,k < tTN < tP,k th X a l do nguyn nhn ngu nhin
Nu tTN>tP,k hoc tTN<-tP.k th X a khng phi l ngu nhin.
3 . X l thng k cc -ng chun.
xc nh -c hm l-ng cht phn tch cn phi da vo -ng
chun biu th mi lin h gia mt quang v nng ca dung dch
cht nghin cu. -ng chun -c x l bng ph-ng php ton hc
thng k nh- sau:
yi l gi tr o -c ca mt quang, xi l gi tr nng ca cht phn
tch.
Yi l gi tr tnh -c t biu thc: Yi = a+b xi
(yi - Yi) l gi tr sai lch ca cc gi tr o.
S trng nhau gia cc gi tr o -c v tnh ton -c tt nht nu
tng cc bnh ph-ng:
n

(yi - Yi)2 = (yi- a- bxi)2 , b nht

1

iu c ngha l:

(yi- a- bxi)2 =-2 (yi- a- bxi) = 0

(yi- a- bxi)2 =-2 (yi- a- bxi) = 0

(yi- a- bxi) = 0

xi(yi- a- bxi)= 0

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yi = na+b xi

xiyi=a xi + b xi2

Gii h ph-ng trnh ny theo nh thc:

A =n xi2- ( xi)2

A1 =

yi xi2 -

A2 =n xiyi -

a =

A1
A

b=

xi xiyi

xi yi

A2
A

Tnh -c a v b, phi nh gi chnh xc ca chng:

Sy2 =

(yi - Yi)2/K =

(yi - Yi)2/(n-1)

Sa2 = Sy2 xi2/ A

Sb2=Sy2.n/ A
chnh xc ca a, b:
a = tP,k.Sa
b=tP,k.Sb

tin cy ca a, b:
a= a a = a tP,k.Sa
b= b b = b tP,k.Sb
Ph-ng trnh -ng chun:
y=(a a)+(b b).x

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Phn II
Thc nghim

I. Ho cht-dng c-my.
1. Ho cht.
Tt c cc ho cht s dng trong lun vn ny u thuc loi tinh
khit ho hc (tkhh) hoc tinh khit phn tch (tkpt). N-c ct 2 ln. Cc
ho cht -c ghi c th trong cc ph-ng php thc nghim dng.
2. Dng c -My mc.
- Dng c thu tinh: Bnh nh mc. Microburet, pipet, bnh tam gic,
cc thu tinh, phu chit...
- Cn phn tch c chnh xc 0,1mg.
- My o pH 420A-Orion.
- My so mu Chroma-model 257.
II. Ph-ng php pha ch cc dung dch dng cho
phn tch.
+ Dung dch 2,6 - diclophenolindophenol 0,001N: Ho tan 0,3000g
2,6 - diclophenolindophenol trong 500ml n-c ct un si, lc dung dch
vo bnh c ngn 1000ml, sau khi ngui thm n-c ct n ngn. Xc
nh chun bng dung dch mui Mohr nh- sau: 5ml dung dch bo ho
natri oxalat thm vo 10ml dung dch indophenol, trn u v nh l-ng
bng dung dch mui Mohr 0,01N cho ti khi mu chuyn t xanh l n
vng; 1ml dung dch mui Mohr t-ng ng vi 10ml dung dch cht mu
0,001N.
+ Dung dch mui Mohr 0,01N: em ho tan 3,9215g mui Mohr vo
500ml n-c ct, thm vo 40ml dung dch axit sunfuric 0,2N v thm n-c
thnh 1000ml.
+ Dung dch bo ho natri oxalat (pha ch bng cch ho tan nng).

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+ Hn hp axit axetic v axit metaphotphoric: Ho tan 15,0000g axit
HPO3 lc vi 40ml axit axetic bng v 200ml n-c ct. Pha long thnh
500ml, lc vo bnh c nt mi v trong t lnh.
+ Dung dch m pH = 4,0: Ho tan 300,00g Natriaxetat trong 700ml
n-c v 1000ml axit axetic bng.
+ Dung dch 2,6 - diclophenolindophenol: Ho tan 25,0mg mui Natri
ca 2,6 - diclophenolindophenol trong n-c nng, lc v pha long vi
n-c ct thnh 200ml.
+ Dung dch axit metaphotphoric: Ho tan lnh 60,00g HPO 3 trong
900ml n-c ct, pha long thnh 1000ml v bo qun trong t lnh.
+ Dung dch axit ascorbic chun: Ho tan 100,0 mg axit ascorbic tinh
khit trong HPO3 6% v pha long thnh 100ml. Ht 25ml dung dch ny
v pha long bng HPO3 6% thnh 1000ml.
III. Xc nh hm l-ng vitamin C bng ph-ng
php chun oxy ho-kh.
1. Nguyn tc ca ph-ng php.
Khi chun oxy ho axit ascorbic bng 2,6 - diclophenolindophenol
trong mi tr-ng axit, axit ascorbic b oxy ho thnh axit dehydroascorbic
v cht mu b kh thun nghch thnh dng khng mu. S kt thc nh
l-ng -c bo hiu bng s xut hin mu hng, chng t s c mt ca
mt l-ng c tha cht mu khng b kh.
y chng ti p dng ph-ng php ny nh l-ng axit ascorbic
trong n-c cam p ng hp nhp t xtrylia. y l nhng mu c cha
cc thnh phn cn tr cng c tnh kh vi cht mu 2,6
diclophenolindophenol trong a s l cc hp cht thuc loi sunfhydryl.
V vy nh l-ng chng ti tin hnh ng thi 2 th nghim song song.
TN1: chun khng c CuSO4 xc nh axit ascorbic v cc cht
kh khc ni chung.
TN2: chun c mt CuSO4 xc nh cc cht kh v cc cht

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sunfhydryl. iu kin ny vitamin C b ph hu ht bi s c mt ca CuSO4.
S chnh lch gia 2 kt qu cho ta l-ng dung dch chun tiu th
t-ng ng vi hm l-ng axit ascorbic c trong mu.
2. Chun b dung dch phn tch.
Ly chnh xc 50ml dung dch n-c cam p ng hp (nhp t xtrylia
c hm l-ng 80,0 mg/1 chai 2 lt) cho vo bnh nh mc 100ml thm hn
hp axit axetic v axit metaphotphoric cho n vch. Lc u, lc v thu
dch lc vitamin C phn tch. Khi hm l-ng vitamin C trong mt ml
dung dch l 0,02mg/1ml.
3. Cch tin hnh.
TN1: Dng pipet ly vo bnh nn dung tch 50ml, 10ml dch lc v
tin hnh chun bng dung dch 2,6 - diclophenolindophenol 0,001N t
microburet cho ti khi xut hin mu hng bn trong 1 pht -c V 1(ml).
TN2: Dng pipet ly vo bnh nn dung tch 50ml, 10ml dch lc thm
vo 1ml dung dch CuSO4 1%, un 10 pht ni cch thu si ph hu ht
vitamin C. ngui v chun bng dung dch 2,6 - diclophenolindophenol
0,001N cho ti khi xut hin mu hng bn trong 1 pht -c V2(ml).
Chun 3 ln ta thu -c kt qu sau.
Bng 1:
m(mg/l)

TT

V1(ml)

V2(ml)

V1- V2 (ml)

2,46

0,40

2,06

0,0181

2,49

0,39

2,10

0,0185

2,52

0,37

2,15

0,0189

vitamin C

0,0185

T s liu thu -c ta tnh -c hm l-ng vitamin C (mg/ml) theo

cng thc sau: m

(V1 V2 ) x0,088
V

Trong : 0,088 l s mg axit ascorbic ng vi 1ml dung dch 2,6 -

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diclophenolindophenol 0,001N.
V: S ml dch lc chun . V = 10ml
4. X l kt qu v tho lun.
Kt qu thu -c chng ti x l theo ph-ng php ton thng k v
thu -c kt qu bng sau:
Bng 2:
Hm l-ng cn

Hm l-ng xc

Hm l-ng

tin cy

xc nh (mg/ml)

nh -c (mg/ml)

X (mg/ml)

q%

.100

0,0181
0,02

0,0185

tTN

X a
SX

|tTN(-2,309)| <
9,9 x 10-4

0,0185

5,35

0,0189

|tp,k (4,30)|
vy sai s
ngu nhin

IV. Xc nh hm l-ng vitamin C bng ph-ng

php o quang.
1. Nguyn tc ca ph-ng php.
Nguyn tc ca ph-ng php l da trn s theo di qu trnh lm mt
mu ca 2,6 - diclophenolindophenol bng dung dch axit ascorbic chun
v bng dch chit.
nh l-ng axit ascorbic trong dch qu, y l nhng mu m dch
chit c mu v c xut hin cc tp cht c tnh kh. Tuy nhin cc tp
cht kh ny phn ng vi 2,6 - diclophenolindophenol chm hn axit
ascorbic do ta c th loi tr nh h-ng ca chng bng cch chit l-ng
tha 2,6 - diclophenolindophenol ch-a b kh trong mi tr-ng phn ng axit
bng xylen. Theo cch lm ny, ta c th ngn tr s tip xc ca dng oxy
ho qu tha ca cht mu vi cc cht tc dng chm tr axit ascorbic.
o mt quang ca l-ng tha cht mu trong xylen. T tnh -c
hm l-ng axit ascorbic c trong mu da vo -ng cong chun xy dng
vi dung dch axit ascorbic chun biu th s ph thuc tuyn tnh gia mt
quang ca l-ng cht mu tha vi nng axit ascorbic chun.

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2. Xc nh cc iu kin ti -u, xy dng -ng chun.
2.1. Chn b-c sng ti -u.
a. Cch tin hnh.
Ht 5ml dung dch axit ascorbic chun pha trn, cho vo bnh
nh mc 25ml, thm 5ml dung dch m axetat, 2ml dung dch cht mu
2,6 - diclophenolindophenol thm n-c ct cho n vch. S xut hin mu
hng ca dung dch chng t s c mt ca mt l-ng tha cht mu khng
b kh trong mi tr-ng axit. Chuyn ton b dung dch trn vo phu chit
dung tch 125ml, sau cho vo 10ml xylen, y phu chit v lc phn
cht mu tha khng b kh vo lp xylen. yn cho 2 t-ng phn lp ri
tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v nh mc n
vch bng xylen. Tin hnh o mt quang cc b-c sng khc nhau
( = 470-710nm) ta thu -c bng sau.
Bng 3: Kho st s ph thuc ca A vo .
Vdd (ml)

Vdd(ml)

Axit ascorbic

cht mu

5,00

(nm)

2,00

430

0,248

5,00

2,00

470

0,323

5,00

2,00

490

0,331

5,00

2,00

520

0,291

5,00

2,00

540

0,177

5,00

2,00

580

0,105

5,00

2,00

600

0,049

5,00

2,00

710

0,007

Mu s

b: Nhn xt.
Gi tr mt quang cao nht v n nh nht l b-c sng

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= 490nm. Do chng ti chn b-c sng ny o mt quang trn
my so mu trong qu trnh thc nghim.
2.2. Kho st pH ti -u.
a. Cch tin hnh:
Chng ti tin hnh nh l-ng vitamin C trong mi tr-ng axt, bi
v trong mi tr-ng axit vitamin C bn vng hn trong mi tr-ng kim.
Kho st nh h-ng ca pH n gi tr mt quang bng cch: ly 6 bnh
nh mc 25ml cho vo mi bnh 5ml dung dch axit ascorbic chun, thm
vo 2ml dung dch cht mu, dng dung dch m v iu chnh pH
khc nhau bng axit HCl v baz NaOH, thm n-c ct vo mi bnh cho
n vch. Cho vo phu chit dung tch 125ml, sau cho vo 10ml xylen,
y phu chit v lc phn cht mu tha khng b kh vo lp xylen.
yn cho 2 t-ng phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh
nh mc 25ml v nh mc n vch bng xylen. Tin hnh o mt
quang trn my so mu ta thu -c kt qu sau:
Bng 4: Kho st s ph thuc ca A vo pH:
Vdd (ml)

Vdd(ml)

Axit ascorbic

cht mu

5,00

Mu s

2,00

pH = 1

0,290

5,00

2,00

pH = 2

0,321

5,00

2,00

pH = 3

0,327

5,00

2,00

pH = 4

0,331

5,00

2,00

pH = 5

0,295

5,00

2,00

pH = 6

0,291

b. Nhn xt:
Gi tr mt quang bn v n nh i vi dung dch c pH = 4. Khi

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dng cc dung dch m ca axit mnh hoc axit yu u dn n s gim
mt quang. Do trong qu trnh thc nghim chng ti chn pH = 4.
2.3. Kho st dung l-ng m cn thit:
a. Cch tin hnh:
Ly 5 bnh nh mc 25ml, cho vo mi bnh 5ml dung dch axit
ascorbic chun, thm vo mi bnh t 2-6 ml dung dch m axetat pH = 4
v 2ml cht mu. Dng n-c ct thm vo mi bnh cho n vch. Cho vo
phu chit dung tch 125ml, sau cho vo 10ml xylen, y phu chit v
lc phn cht mu tha khng b kh vo lp xylen. yn cho 2 t-ng
phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v
nh mc n vch bng xylen. Tin hnh o mt quang trn my so
mu ta thu -c kt qu sau:
Bng 5:
Vdd (ml)

Vdd(ml)

Vdd(ml) m

Axit ascorbic

cht mu

PH = 4

5,00

2,00

2,00

0,276

5,00

2,00

3,00

0,292

5,00

2,00

4,00

0,310

5,00

2,00

5,00

0,331

5,00

2,00

6,00

0,302

Mu s

b. Nhn xt: Gi tr mt quang n nh v t cc i khi l-ng

dung dch m l 5ml. Do l-ng dung dch m thch hp dng trong
qu trnh thc nghim l 5ml ng vi 5ml dung dch axit ascorbic chun.
2.4. Kho st l-ng thuc th thch hp:
a. Cch tin hnh:
Cho vo 5 bnh nh mc 25ml, mi bnh 5ml dung dch axit ascorbic
chun, thm vo 5ml dung dch m axetat pH = 4, ln l-t thm dung dch
cht mu vi th tch khc nhau v nh mc n vch bng n-c ct. Cho

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vo phu chit dung tch 125ml, sau cho vo 10ml xylen, y phu chit
v lc phn cht mu tha khng b kh vo lp xylen. yn cho 2 t-ng
phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v
nh mc n vch bng xylen. Tin hnh o mt quang trn my so
mu ta thu -c kt qu sau:
Bng 6:
Vdd (ml)

Vdd(ml)

Vdd(ml) m

Axit ascorbic

cht mu

pH = 4

5,00

0,50

5,00

0,000

5,00

1,00

5,00

0,082

5,00

1,50

5,00

0,164

5,00

2,00

5,00

0,331

5,00

2,50

5,00

0,651

Mu s

b. Nhn xt: L-ng thuc th cng d- th gi tr mt quang cng

cao. Tuy nhin l-ng thuc th d- nhiu hoc qu t th u dn n sai s
ln, do trong qu trnh thc nghim chng ti chn l-ng thuc th
thch hp l 2ml.
2.5. Kho st l-ng dung mi thch hp.
a. Cch tin hnh:
Ly 5 bnh nh mc 25ml, cho vo mi bnh 5ml dung dch axit
ascorbic chun, thm vo mi bnh 5ml dung dch m, 2ml cht mu, v
thm n-c ct cho n vch. Cho vo phu chit dung tch 125ml, sau
cho vo mi phu t 6 n 12 ml xylen, y phu chit v lc phn cht
mu tha khng b kh vo lp xylen. yn cho 2 t-ng phn lp, ri tin
hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v nh mc n
vch bng xylen. Tin hnh o mt quang trn my so mu ta thu -c
kt qu sau:

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Bng 7:
Vdd (ml)

Vdd(ml)

Axit ascorbic

cht mu

5,00

Mu s

V(ml) xylen

2,00

6,00

0,318

5,00

2,00

8,00

0,329

5,00

2,00

10,00

0,331

5,00

2,00

11,00

0,330

5,00

2,00

12,00

0,322

b. Nhn xt: Gi tr mt quang t cc i khi l-ng dung mi l

10ml. Do l-ng dung mi thch hp dng chit l-ng tha cht mu
trong qu trnh thc nghim chng ti chn l 10ml.
2.5. Kho st thi gian nh h-ng n mt quang.
a. Cch tin hnh:
Cho vo bnh nh mc 25ml, 5ml dung dch axit ascorbic chun,
thm vo 5ml dung dch m axetat pH = 4; 2ml dung dch cht mu. Dng
n-c ct thm vo mi bnh cho n vch. Chuyn dung dch mi bnh
vo phu chit dung tch 125ml. Sau cho vo 10ml xylen, y phu chit
v lc phn cht mu tha khng b kh vo lp xylen. yn cho 2 t-ng
phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v
nh mc n vch bng xylen. Tin hnh o ngay mt quang trn my
so mu. Sau o gi tr mt quang cc mc thi gian khc nhau ta
thu -c kt qu bng sau:

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Bng 8: Kho st s ph thuc ca A vo thi gian o
Vdd (ml)

Vdd(ml)

Thi gian o

Axit ascorbic

cht mu

(pht)

5,00

2,00

0,329

5,00

2,00

0,331

5,00

2,00

10

0,331

5,00

2,00

20

0,331

5,00

2,00

30

0,331

5,00

2,00

40

0,331

5,00

2,00

50

0,331

5,00

2,00

60

0,330

Mu s

b. Nhn xt: Trong khong thi gian t 5-50 pht khng c s mt

mt no v gi tr mt quang. Nh- vy mu trong mi tr-ng xylen kh
bn vng, cho nn khng cn o vi vng, quan trng l lc lin tc t mt
th tch nh- nhau khi tin hnh chit l-ng tha cht mu vo lp xylen.
2.6. Xy dng -ng chun.
a. Cch tin hnh:
Cho vo 9 bnh nh mc 25ml ln l-t t 1-12ml dung dch axit
ascorbic chun, thm vo mi bnh 5ml dung dch m axetat pH = 4; 2ml
dung dch cht mu, v nh mc n vch bng n-c ct. Cho vo phu
chit dung tch 125ml, sau cho vo 10ml xylen, y phu chit v lc
phn cht mu tha khng b kh vo lp xylen. yn cho 2 t-ng phn
lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v nh
mc n vch bng xylen. Tin hnh o mt quang trn my so mu ta
thu -c kt qu bng sau:

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Bng 9: Kho st s ph thuc ca A vo hm l-ng dung dch chun

Vdd (ml)

Vdd (ml)

Hm l-ng

Axit ascorbic

Cht mu

(mg)axit ascorbic

1,00

2,00

0,0250

0,390

2,00

2,00

0,0500

0,375

3,00

2,00

0,0750

0,361

4,00

2,00

0,1000

0,345

5,00

2,00

0,1250

0,331

6,00

2,00

0,1500

0,317

8,00

2,00

0,2000

0,289

10,00

2,00

0,2500

0,260

12,00

2,00

0,3000

0,228

Mu s

b. Nhn xt: Khi hm l-ng ascorbic ln hn 0,25mg th gi tr mt

quang khng thun theo nh lut Bughe-Lambe-Bia.
T gi tr thu -c bng trn ta xy dng th biu th s ph thuc
ca mt quang vo hm l-ng axit ascorbic nh- sau:
A
0,400

0,45
0,4
0,350
0,35
0,3
0,300
0,25

East

0,2
0,250
0,15
0,1
0,200
0,05
0
0,05 0,10 0,15
0,03 0,08 0,13

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0,2
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3. Xc nh hm l-ng vitamin C trong mu t to bng ph-ng
php o quang.
c c s chc chn, khoa hc p dng ph-ng php phn tch v
cc iu kin ti -u chn. Tr-c khi phn tch hm l-ng vitamin C
trong mu n-c p tri cy ng hp v hoa qu, th chng ti th nghim
xc nh hm l-ng vitamin C trong mu t to.
3.1. Chun b mu:
Cn chnh xc trn cn phn tch 25,0 mg axit ascorbic tinh khit v
ho tan trong HPO3 6%, pha long thnh 1000ml. Dung dch thu -c c
hm l-ng vitamin C l 0,025 mg/ml.
3.2. Cch tin hnh:
Ht 5ml dung dch axit ascorbic chun pha trn cho vo bnh nh
mc 25ml. Thm 5ml dung dch m axetat pH = 4; 2ml dung dch cht
mu, v nh mc n vch bng n-c ct. Cho vo phu chit dung tch
125ml, sau cho vo 10ml xylen, y phu chit v lc phn cht mu
tha khng b kh vo lp xylen. yn cho 2 t-ng phn lp, ri tin
hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v nh mc n
vch bng xylen. Tin hnh o mt quang b-c sng = 490nm. Ta
thu -c kt qu bng sau:
Bng 10:

Hm l-ng xc

Hm l-ng

tin cy

TT
nh -c (mg/ml)

24,94 . 10-3

24,92 . 10-3

q%

.100

t(TN)

|t(TN)(-0,333)|

24,92. 10-3

0,035.10-3

0,14

<

|tb,k(4,30)|

24,90 . 10-3
S.S ngu nhin
Bng ph-ng php ton hc thng k chng ti tin hnh so snh

cc gi tr xc nh -c vi gi tr cho tr-c (a = 25 . 10 -3). S sai khc

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gia X v a l do nguyn nhn ngu nhin. iu cho php p dng
ph-ng php o quang phn tch hm l-ng vitamin C trong dch qu
vi cc iu kin ti -u nhn -c v dung mi, pH v b-c sng
4. Tin hnh xc nh hm l-ng vitamin C trong n-c p tri cy
nguyn cht ng hp v mt s hoa qu c bn trn th tr-ng thnh
ph Vinh.
4.1. Xc nh hm l-ng vitamin C trong mt s n-c p tri cy
nguyn cht ng hp (nhp t xtrylia).
4.1.1. X l mu:
Ly chnh xc 50ml n-c cam p nguyn cht ng hp (nhp t
xtrylia) vi hm l-ng vitamin C ghi trn nhn l 80mg/1 chai 2 lt. Cho
vo bnh nh mc 100ml v nh mc n vch bng HPO 3 6% lc u v
lc dch trn phu thu tinh xp.
4.1.2. Kho st cc yu t nh h-ng n qu trnh nh l-ng bng
ph-ng php chit indophenol-xylen.
a. nh h-ng do dch chit c mu:
Nhn thy dch chit thu -c c mu. V vy nh l-ng vitamn C
bng ph-ng php chit indophenol-xylen, th tr-c ht phi xem dch lc
c mu c chit -c bng xylen khng.
Th nghim bng cch trn 5ml dch chit axit thu -c trn vi 5ml
m v chit vi 10ml xylen. Chuyn lp xylen vo cuvt v tin hnh o
mt quang ta thu -c gi tr D ~ 0. Nh- vy dch chit c mu khng
chit -c bng xylen, do ta c th p dng ph-ng php ny nh
l-ng vitamin C trong mt s dch qu c mu.
b. nh h-ng ca mt s tp cht kh:
Khi nh l-ng vitamin C trong dch qu th-ng gp cc cht cn tr.
Bng ph-ng php sc k giy vi h dung mi l: n-butanol-axit axeticn-c (4:1:5) v pht hin bng dung dch 2,6-diclophenolindophenol.

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Ng-i ta xc nh -c cc thnh phn cn tr l cc hp cht
thuc loi sunfhydrin v cc cht c tnh kh khc. y l nhng tp cht
cng c tnh kh vi cht mu 2,6-diclophenolindophenol. Tuy nhin s
mt mu do axit ascorbic thc t xy ra tc thi, cn s mt mu do cc tp
cht c tnh kh phn nhiu chm hn. Do c th loi tr nh h-ng ca
cc cht kh ny bng cch chit l-ng tha cht mu 2,6 iclophenolinophenol ch-a b kh trong mi tr-ng phn ng axit bng
xylen.
Vy thi gian thch hp chit l-ng tha cht mu vo mi tr-ng
xylen l bao nhiu? trong khong thi gian cc tp cht kh ch-a
phn ng vi 2,6-diclophenolindophenol.
c. Kho st khong thi gian thch hp chit l-ng tha cht mu
vo mi tr-ng xylen, khi cc tp cht kh ch-a phn ng vi 2,6diclophenolindophenol.
Tin hnh bng cch: Ht 5ml dch qu thu -c trn cho vo bnh
nh mc 25ml, thm 5ml dung dch m, 2ml cht mu v thm n-c cho
n vch. Tin hnh o ngay mt quang ca dung dch trn my so mu,
sau o gi tr mt quang cc mc thi gian khc nhau ta thu -c
kt qu bng sau:
Bng 11: Kho st s ph thuc ca l-ng cht mu tha vo thi gian.
Vdd (ml)

Vdd (ml)

Thi gian

Axit ascorbic

Cht mu

(giy)

5,00

2,00

0,330

5,00

2,00

10

0,331

5,00

2,00

20

0,331

5,00

2,00

40

0,331

5,00

2,00

60

0,331

5,00

2,00

80

0,325

Ln o

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Nhn xt: T 10-60 giy gi tr mt quang khng thay i. Sau 1
pht gi tr mt quang bt u gim xung do cc tp cht kh bt
u phn ng vi 2,6-diclophenolindophenol. Nh- vy khong thi gian
thch hp nht chit l-ng tha 2,6-diclophenolindophenol ch-a b kh
trong mi tr-ng phn ng axit bng xylen l t 10-60 giy.
4.1.3..Tin hnh nh l-ng.
Cho vo 3 bnh nh mc 25ml, mi bnh 5ml dch lc trn thm vo
5ml dung dch m axetat pH = 4 v 2ml dung dch cht mu 2,6 diclophenolindophenol, thm n-c ct cho n vch. Cho nhanh vo phu
chit dung tch 125ml, sau cho vo 10ml xylen, y phu chit v lc
trong khong thi gian t 15-20 giy (cn gia ng thi gian chit trong
khong 1 pht loi tr nh h-ng ca cc thnh phn kh khc c phn
ng vi cht mu nh-ng chm hn axit ascorbic). yn cho 2 t-ng phn
lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc 25ml v nh
mc n vch bng xylen. Tin hnh o mt quang b-c sng =
490nm. Ta thu -c kt qu bng sau:
Bng 12:

Ln

Mu

tin cy

vitamin C

(mg/ml)

A
o

N-c

m(mg/ml)

q%

0,346

0,0201

0,347

0,0198

0,348

0,0195

.100

tTN

X a
SX

|tTN(-1.154)| <

cam
0,0198

7,44x10-4

3,75

|tp,k (4,30)| sai s

p ng hp
ngu nhin

4.1.4. X l kt qu v tho lun:

Bng ph-ng php ton hc thng k chng ti tin hnh so snh gi
tr xc nh -c vi gi tr cho tr-c (-c coi l gi tr chun a,
a = 0,02mg/ml). S sai khc gia m v axit l do nguyn nhn ngu nhin.

L Th Sen

40

Lp: 41B2

Kho lun tt nghip

Vy chng ta c th p dng ph-ng php ny xc nh hm l-ng
vitamin C trong n-c p tri cy nguyn cht ng hp.
4.2. Xc nh hm l-ng vitamin C trong mt s loi qu.
4.2.1. X l mu:
Nghin nh nhanh chng mt l-ng qu cn th (cam H-ng Sn,
cam Ngha n) trong mt ci s mi tr-ng axit HPO 3 6%. Cn 10g bt
nghin chuyn vo bnh nh mc 100ml v lm y bng HPO 3 6%. Lc
dch trn phu thu tinh xp.
4.2.2. Tin hnh nh l-ng.
a. Mu cam trong ngy u:
Cho vo 3 bnh nh mc 25ml, mi bnh 5ml dch cam H-ng Sn
mi pha trn, thm vo 5ml dung dch m axetat pH = 4 v 2ml dung
dch cht mu 2,6-diclophenolindophenol. thm n-c ct cho n vch. Cho
nhanh vo phu chit dung tch 125ml, sau cho vo 10ml xylen, y
phu chit v lc trong khong thi gian t 15-20 giy (cn gi ng thi
gian chit trong khong 1 pht loi tr nh h-ng ca cc thnh phn
kh khc c phn ng vi cht mu nh-ng chm hn axit ascorbic). yn
cho 2 t-ng phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh
mc 25ml v nh mc n vch bng xylen. Tin hnh o mt quang
b-c sng = 490nm. Ta thu -c kt qu bng (13)
Lm th nghim t-ng t nh- trn vi dch cam Ngha n ta thu -c
gi tr mt quang ( bng 14).
b. Mu cam 1 ngy sau ph mu:
Cho vo 3 bnh nh mc 25ml, mi bnh 5ml dch cam H-ng Sn
qua mt ngy x l, thm vo 5ml dung dch m axetat pH = 4, 2ml
dung dch cht mu 2,6-diclophenolindophenol, thm n-c ct cho n vch.
Cho nhanh vo phu chit dung tch 125ml, sau cho vo 10ml xylen, y
phu chit v lc trong khong thi gian t 15-20 giy (cn gia ng thi
gian chit trong khong 1 pht loi tr nh h-ng ca cc thnh phn kh

L Th Sen

41

Lp: 41B2

Kho lun tt nghip

khc c phn ng vi cht mu nh-ng chm hn axit ascorbic). yn cho 2
t-ng phn lp, ri tin hnh chit. Chuyn lp xylen vo bnh nh mc
25ml v nh mc n vch bng xylen. Tin hnh o mt quang ph
b-c sng = 490nm. Ta thu -c kt qu bng (13).
Lm th nghim t-ng t nh- trn vi dch cam Ngha n ta thu -c
gi tr mt quang ( bng 14).
Bng 13:
Mu s

Ngy u

Sau 1 ngy

Vdd (ml)

Vdd (ml)

Cam

Cht mu

5,00

2,00

0,289

5,00

2,00

0,288

5,00

2,00

0,286

5,00

2,00

0,349

5,00

2,00

0,349

5,00

2,00

0,348

Ln o

Bng 14:
Mu s

Ngy u

Sau 1 ngy

L Th Sen

Vdd (ml)

Vdd (ml)

Cam

Cht mu

5,00

2,00

0,313

5,00

2,00

0,314

5,00

2,00

0,311

5,00

2,00

0,351

5,00

2,00

0,349

5,00

2,00

0,352

Ln o

42

Lp: 41B2

Kho lun tt nghip

4.1.3. X l kt qu v tho lun.
a. X l kt qu.
Mu cam H-ng Sn:
Bng 15: Kt qu phn tch hm l-ng vitamin C trong mu cam
H-ng Sn.

Mu s

Ngy u

Sau 1 ngy

m(mg/100g)

Ln o

0,289

39,9800

0,288

40,0000

0,287

40,0800

0,349

19,0600

0,349

19,0600

0,348

19,0900

vitamin C

40,02

0,1310

19,07

0,0430

Bng 16: Kt qu phn tch hm l-ng vitamin C trong Mu cam

Ngha n
Mu s

Ngy u

Sau 1 ngy

m(mg/100g)

Ln o

0,313

31,2500

0,314

31,2400

0,311

31,2900

0,351

18,4500

0,350

18,4200

0,352

18,3600

vitamin C

31,2600

0,065

18,4100

0,114

Sau khi o mt quang, t cc gi tr thu -c ta -a ln -ng

chun, t suy ra nng axit ascorbic trong dch cam. Bit nng ta
tnh -c hm l-ng vitamin C trong tng mu. Kt qu thu -c chng ti

L Th Sen

43

Lp: 41B2

Kho lun tt nghip

x l theo ph-ng php ton thng k .
b. Tho lun kt qu:
T kt qu phn tch -c chng ti thy hm l-ng vitamin C trong
mu cam H-ng Sn nhiu hn hm l-ng vitamin C trong mu cam Ngha
n v hm l-ng vitamin C gim i rt nhiu nu mu cam ph mu
qua mt ngy nhit phng. iu chng t rng dung dch axit
ascorbic d b oxy ho trong khng kh.

L Th Sen

44

Lp: 41B2

Kho lun tt nghip

phn III
Kt lun
1. tng quan v tm quan trng ca vitamin C, cc ngun vitamin
C trong thin nhin, tnh cht ca vitamin C, cc ph-ng php nh
l-ng vitamin C.
2. chn -c thuc th thch hp c kh nng tham gia phn ng
oxy ho-kh thun nghch vi axit ascorbic l 2,6-diclophenolindophenol
v xc nh -c cc i hp th ca cht mu ny b-c sng bng
490nm.
3. loi tr -c nh h-ng ca cc cht cn tr trong qu trnh nh
l-ng bng cch chit l-ng tha ca cht mu 2,6- diclophenolindophenol
trong mi tr-ng phn ng bng xylen. Thi gian chit thch hp nht l
trong 1pht (tnh t lc bt u cho cht mu vo dung dch phn tch).
4. tin hnh xc nh -c hm l-ng vitamin C trong n-c cam p
nguyn cht ng hp (nhp t xtrylia) v cam (H-ng Sn, Ngha n)
c bn trn th tr-ng thnh ph Vinh hin nay. V thu -c kt qu nhsau:
- Hm l-ng vitamin C trong mt chai n-c cam p nguyn cht ng
hp:
Theo ph-ng php chun oxy ho l 74 1,972 (mg).
Theo ph-ng php chun trc quang l 79,2 2,976 (mg).
- Hm l-ng vitamin C trong trong mu cam:
Phn tch ngy u:

L Th Sen

+ Cam H-ng Sn:

40,02 0,131 (mg /100g)

+ Cam Ngha n:

31,26 0,065 (mg /100g)

45

Lp: 41B2

Kho lun tt nghip

Phn tch sau mt ngy ph mu:
+ Cam H-ng Sn:

19,07 0,043 (mg /100g)

+ Cam Ngha n:

18,41 0,114 (mg /100g)

5. Do thi gian khng cho php nn chng ti khng th tin hnh

phn tch hm l-ng vitamin C khi dng vi cc thuc th khc nhau, ch-a
xc nh -c hm l-ng vitamin C trong mt s hoa qu khc. Chng ti
hy vng quy trnh phn tch ny c th -c p dng phn tch hm
l-ng vitamin C trong cc loi hoa qu khc nhau v c bit c th s
dng kim nghim hm l-ng vitamin C trong mt s loi n-c p tri
cy nguyn cht (ni nhp, ngoi nhp ) ang c bn trn th tr-ng hin
nay.
V iu kin c hn nn ng-i thc hin ti khng trnh khi nhng
thiu st. Rt mong -c s gp v ch bo ca cc thy c gio v cc
bn sinh vin.

L Th Sen

46

Lp: 41B2

Kho lun tt nghip

Ti liu tham kho

1. L Vn Tri, Nguyn Ngc Don. Sinh hc vitamin. NXB KHKT, H

Ni. 1987
2. L Don Din, l Duy Thu, M Xuyn, Hong Vn Tuyn. Vitamin v
i sng. NXB KHTK, H Ni.1978
3. L Ngc T, Phm Quc Thng, L Don Din, Bi c Hi, La Vn
Ch, Nguyn Th Thnh. Ho sinh hc cng nghip. NXB i hc &
THCN, H Ni 1974
4. Trn Th Hong Ba, V Th By, Nguyn Kim Chi, Don Huy Khc,
Nguyn Quang Lun. nh l-ng vitamin. NXB Y hc H Ni, 1976
5. ng Th Thu, Nguyn Th Xun Sm, T Kim Anh. Th nghim ho
sinh cng nghip. Tr-ng H Bch khoa H Ni, H ni 1997
6. Phm Th Trn Chu, Trn Th ng. Ho sinh hc. NXB Gio dc, H
Ni 1999
7. Nguyn Hu Cht. Enzym v xc tc sinh hc. NXB Y hc H Ni,
1983
8. Trn ch. Ho sinh hc. NXB Gio dc, H Ni
9. H Vit Qu. Phn tch l ho. NXB Gio dc, H Ni 2000
10. Nguyn Tinh Dung. Ho phn tch, T1, T2. NXB Gio dc, H Ni
1991
11. Nguyn Khc Ngha. p dng ton hc thng k x l s liu thc
nghim. H Vinh

L Th Sen

47

Lp: 41B2

Kho lun tt nghip

Mc lc
Trang
Li m u

Phn i. Tng quan ti liu

I. Gii thiu v vitamin C

1. L-c s

2. Vai tr ca vitamin C trong i sng con ng-i

3. Cc ngun vitamin C trong thin nhin

4. Cng thc ho hc

5. Ph-ng php iu ch

6. Tnh cht ca vitamin C

II Cc ph-ng php xc nh vitamin C

10

1. Nhn xt chung

10

2. nh l-ng axit ascorbic bng ph-ng php vt l

11

2.1. Ph-ng php quang ph

11

2.2. Ph-ng php cc ph

11

2.3. Ph-ng php sc k

14

3. nh l-ng axit ascorbic bng ph-ng php ho hc

16

3.1. nh l-ng bng ph-ng php oxyho

16

3.2. nh l-ng axit ascorbic bng ph-ng php o quang

19

III. c im ca thuc th hu c 2,6 - diclophenolindophenol

20

IV. Kt lun

21

V. X l thng k cc s liu thc nghim

22
26

Phn ii: Thc nghim

I. Ho cht, dng c, my mc

26

II. Ph-ng php pha ch cc dung dch dng cho phn tch

26

III. Xc nh hm l-ng vitamin C bng ph-ng php chun oxy

ho kh

L Th Sen

48

Lp: 41B2

27

Kho lun tt nghip

1. Nguyn tc ca ph-ng php

27

2. Chun b dung dch phn tch

28

3. Cch tin hnh

28

4. X l kt qu v tho lun

29

IV. Xc nh hm l-ng vitamin C bng ph-ng php o quang

29

1. Nguyn tc ca ph-ng php

29

2. Xc nh cc iu kin ti -u, xy dng -ng chun

30

3. Xc nh hm l-ng vitamin C trong mu t to bng ph-ng

php o quang

36

4. Xc nh hm l-ng vitamin C trong n-c p tri cy nguyn cht

38

ng hp v mt s hoa qu c bn trn th tr-ng thanh ph Vinh

45

Phn iii: Kt lun

47

Ti liu tham kho

L Th Sen

49

Lp: 41B2