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Of Ascorbic Acid
Introduction
based on oxidation-reduction.
low pH. This method is widely used and it only measures the
Kuether, 1943) t
with 10 per cent HC1 (iron free) washed several times with
and by this procedure one can measure both the reduced ascorbi
methods.
Experimental
Indophenol Dye.
indophenol dye.
4. 2,6-Dichlorophenolindophenol
distilled water.
Reaction Mechanism s
0=C
I
HO--C
II
HO-C
i
H-C
I
HO-C-H
ch2oh
2,6-dichlorophenolindophenol
(oxidised form,blue or pink)
Ascorbic Acid
CHgOH
2,6-dichlorophenolindophenol
(reduced form - colorless)
Dehydroascorbic Acid
In Tissue Homogenates s
of the tube, until the blue dye turned colorless. The value
Principle i-
Reaction Mechanism s
H H
0=C-.
0=i]
J, 0
0—0 ■
1 I + 0 2 H20
H- C—’
I
HO-C-H
™2°H 2,4-dinitrophenyl- NO 1
2 HO-C-H
hydrazine I
CH20H
Dehydroascorbie
Acid
Bis-2,4-dinitrophenylhydrazine
derivative
of glacial acetic acid was added and the volume was made upto
solution
acid.
longer showed a test for chloride and for iron. It was then
bottle
,53
solution
7* 9N H2SG4
Procedure :
norit method s
Bromine Method :
Reagents : 1. Bromine
Procedure ;
tube (80 ml) from the excess liquid bromime, and a current
Indophenoloxidation method
hydrazine.
Reagents t
1. 2,6-dichlorophenolindophenol
in norit method.
Procedure i
acetic acid solution and the other procedure was the same
was added in different amounts (40, 50, 60, 80, 100 yag
Results s
methods.
TABLE - I
1 WJ 9
vation
I Norit
j Charcoal |Bromine
’I Indophenol Method
J Method
| Method
i Reduced Total j Total Total
ascorbic jascorbie 1 ascorbic
I I ascorbic
acid acid ! acid I acid
1a Liver 59 60 59 66
1b II
53 52 52 58
1c If 49 50 49 55
2a Kidney 40 41 40 47
2b n
37 36 36 42
ii
2c 37 35 35 43
3a Brain 82 80 81 87
3b II
79. 80 78 84
it
3c 79 79 77 85
4a Intestine 64 62 63 69
4b II
59 61 58 65
4c II
70 65 68 74
TABLE - II
Recovery of ascorbic acid added to different tissue (liver, kidney and intestine)homogenates
of rat estimated by three different methods.
A s c o rb ic A s c o r b i c A c id r e c o v e r e d ( y u g /m l t i s s u e h o m o g e n a te )
a c id N o rit c h a rc o a l
N o. of added I n d o p h e n o l M e th o d M e th o d B r o m in e M e th o d
o b s e r T is s u e ( /u g /m l
v a tio n T o ta l P e r c e n t ' a Ts coot ar bl i c P e r c e n t
' t i s s u e Ra se cd ou cr be di c a Ts coot ar bl i c P e r c e n t
re c o v e ry a s c o r b i c re c o v e ry
hom oge a c id a c id a c id a c id re c o v e ry
i
n a te )
i
i
i
i
i
ymm
m
L iv e r 50
s
O
-4
i—
r*
•
r** CO
o o
1>
86
CM
m 0\
m
97 97
s
T*
s
a\ vo
no VO
itk r» •4 VO
•4
00 VO 00
00 N0 CM o
ON VO
v~
00
VO
t—
•4
»—
• •
t—
.4
ON ON ON
OV ON
cn •4
r>-
100 101
I
1
1
i
i
1
CM
CM
ON
f"
cn
K id n e y 33 37
c
•
o o
*4
CM
77 68 79 1 0 2 .5
s
00
00 00
•
f> 00
On
06 O ON
101
cn
6o 88
s
m 00 CM
O
•
*•*
m in VO
VO
o
•
00 VO• .4•
T“
o
On ON ON
C\ On ON
<a
80 109 119
1
I
s
o
vo
-
in
In te s tin e 0 68 75
s
CM
o
•
CM
CM
>n
in
113 9 5 .7 117 129
=
r— O
CM
ON
ON
On
»n
80
s
r*
cn On -4
vo
VO 00
•
vo
r- T“
•
T-* O
r-
in
ON ON On
ON cn
cn
cn -3-
cn On
ON in
-4 in
100 97
61
H E A D IN G
KLETT
0 10 20 30 40
ASCORBIC ACID IN jJi g
FIG. 1
Discussion i
Summary s
5?