Investigations on the determination of the crude fiber content of some feed materials
by James L Milne
A THESIS Submitted to the Graduate Faculty in partial fulfillment of the requirements for the degree
of Master of Science in Chemistry
Montana State University
© Copyright by James L Milne (1951)
Abstract:
Certain modifications of the present official crude fiber determinations have been investigated.
Particular emphasis has been given to the effects of varying temperature and pressure on the hydrolytic
processes involved.
The effectiveness of various hydrolytic agents used singly or in combinations was studied. The
comparative results of the modifications are discussed.
INVESTIGATIONS ON THE DETERMINATION QF
THE- CHDDE FIBER CONTENT OF SOME FEED MATERIALS
by
JAMES L0 MILNE
A THESIS
S ub m itted ' to th e G raduate F a c u lty
in
p a r t i a l f u l f i l l m e n t o f th e re q u ire m e n ts
f o r th e d eg ree o f
M aster o f S cien ce i n C hem istry
Montana S ta te C o lleg e
'Approved?-
Bozeman, Montana
J u ly , 1951 ■
/I/ 6
—2 —
TABLE OF CONTENTS
S e c tio n
Page
I.
A b s tra c t . . . . . . . . . . . . . . . . . . . . .
3
In tro d u ctio n o . . . . . . . . . . . .
I4.
XX.
III.
E x p e rim en ta l P ro ced u re . . . . . . .
. . . . . .
............................
IV 0 E x p erim en tal R e s u lts . . . . . . . . . . . . . . .
Vc Dis c u ss io n . . . . . . . . . . . . . . . .
V I.
V II.
5
9
. . . • •
13
. . . . . . .
23
B ib lio g ra p h y . . . . . . . . . . . . . . . . . . .
2 It
Summary . . . o . . . . . . . . . . .
103823
™a
I.
^
CO
ABSTRACT
C e r ta in m o d ific a tio n s o f th e p r e s e n t o f f i c i a l crude f i b e r d eterm in a­
tio n s have b een in v e s t i g a t e d . P a r t i c u l a r em phasis has b een g iv e n t o th e
e f f e c t s o f .v a r y in g te m p e ra tu re and p r e s s u r e on th e h y d r o ly tic p ro c e s s e s
in v o lv e d .
The e f f e c t iv e n e s s o f v a rio u s h y d r o ly t ic a g e n ts used s in g ly o r i n com­
b in a tio n s was s tu d ie d . The com parative r e s u l t s o f th e m o d ific a tio n s a re
d is c u s s e d .
“ 1(. “
II0
INTRODUCTION
'
' '
The te rm crude f i b e r as u sed i n t h i s t h e s i s i s d e fin e d as th e o rg a n ic
m a te r ia l which remains, in s o lu b le / a f t e r a fe e d m a te r ia l has b een e x tr a c te d
.with d i - e t h y l ethfer and s u b je c te d to th e a c ti o n o f h y d r o ly tic a g e n ts under
s t a t e d c o n d itio n s o f tim e , te m p e ra tu re , and c o n c e n tr a tio n s .
The f e s u l t e - b f -rec en t—c o lIa B o ra tiv e rs.tudy -o n -feed a n a ly s e s c a r r ie d
o u t .b y s e v e r a l s t a t e A g r ic u ltu r a l Experim ent S ta tio n s in d ic a te d t h a t a wide
v a r i a t i o n i n th e .crude f i b e r c o n te n t o f a fe e d .sa m p le could be e x p e c te d .
I t" was n o te d from, .th is c o l l e c t i v e s tu d y t h a t th o s e E xperim ent S ta tio n s
s itu a te d , in . th e h ig h e r e le v a tio n s , o f the- c o u n try .u stU illy r e p o r te d h ig h e r
crude f i b e r v a lu e s th a n th o s e Experim ent S ta tio n s w hich.w ere lo c a te d in .
.th e a re a s, of...lower, e le v a tio n .
fo ld .
.The p u rp o se o f . . t h i s . i n v e s t i g a t i o n was two­
The. f i r s t problem o f i n t e r e s t , concerned th e p o s s ib le e f f e c t s o f
e le v a te d te m p e ra tu re s and p r e s s u r e on th e h y d r o l i t i c r e a c tio n s in v o lv e d
i n th e crude f i b e r d e te r m in a tio n s .
th e p r e s e n t a n a ly s e s
I t was hoped t h a t th e s e v a r ia b le s in '
could be e f f e c t i v e l y s ta n d a rd iz e d so t h a t th e v a r­
i a t i o n s i n th e r e s u l t s b ro u g h t ab o u t b y th e d if f e r e n c e i n a l t i t u d e s
could be e lim in a te d .
The second p u rp o se o f i n v e s t i g a t i o n was to es­
t a b l i s h th e e f f i c i e n c y o f v a rio u s hyd p l y t i c a g e n ts which c o u ld be u sed
'I "
s in g ly o r i n com binations 0
.
III.
A.
EXPERIMENTAL PROCEDURE
M a t e r i a l s .Four d i f f e r e n t liv e s to c k fe e d s were s e le c te d as th e b a s ic m a te r ia ls
fo r th is in v e s tig a tio n .
These fe e d s were I , o a t hay, 2 , a s ta n d a rd m ix tu re
d a ir y fe e d c o n ta in in g soy bean m eal, 3, a l f a l f a hay, and U, b a r le y .
Feeds
No. I and No. 3 were c o n sid e re d r e p r e s e n ta tiv e o f fe e d m a te r ia ls having a
high crude f i b e r c o n te n t w h ile fe e d s No. 2 and No. U were s e le c te d because
o f t h e i r r e l a t i v e l y low crude f i b e r p e rc e n ta g e .
The fe e d s were a i r - d r i e d
and ground i n a W iley M ill to p a ss th ro u g h a LO-mesh s c r e e n .
A fte r g rin d ­
in g , th e sam ples were a i r d r ie d and s to r e d i n t i g h t l y c lo se d g la s s j a r s .
B.
Crude F ib e r D e te rm in a tio n s Using O f f i c i a l M ethod.S e v e ra l crude f i b e r d e te rm in a tio n s were made on each fe e d m a te r ia l
a c c o rd in g to th e O f f i c i a l and T e n ta tiv e Methods o f A n a ly sis o f A.O.A.C.
( lU ) .
The r e s u l t s o f th e s e a n a ly s e s a re ta b u la te d i n T able I .
TABLE I
S ta n d a rd Crude F ib e r P e rc e n ta g e s
O b tained by th e O f f i c i a l Methods
o f A n a ly sis
Feed
No. o f
T e s ts
I
2
3
b
6
6
6
6
■
Average
P e rc e n tag e
H ig h est
P e rcen tag e
2 6 .S 6
7,it3
23.51
3 .5 6
27.85
7.5 5
26.12
3 .8 5
Lowest
P e rcen tag e
25.10
6.30
25.00
3.o5
I t w i l l be no ted T able I c o n ta in s th e in fo rm a tio n c o n cern in g th e
h ig h e s t, th e lo w e st, and th e av erag e o f th e d e te rm in a tio n s f o r each
fe e d
6 “
Ce
Crude F ib e r D e te rm in a tio n s by E le v a tin g Tem perature and P r e s s u r e ,T q o r d e r to c o n tr o l th e te m p e ra tu re and p r e s s u r e , th e h y d r o ly tic re a c ­
t i o n s were conducted i n a s ta n d a rd la b o r a to r y a u to c la v e «
I,
A u to c la v e o-
The a u to c la v e was equipped w ith th e u s u a l p re s s u re
gauge and a th erm o m eter0
F u rth erm o re, t h i s p a r t i c u l a r a u to c la v e w as-so
c o n s tru c tu e d t h a t th e p r e s s u r e could b e v a r ie d a t w i l l by means o f an ad­
j u s t a b l e diaphragm o
2 o C o n ta in e r s omeyer f l a s k s e
The d e te rm in a tio n s were c a r r ie d o u t i n £00 ml E r le n -
I n each c a se , e x c e p t where o th e rw is e n o te d , £0 ml o f th e
h y d r o ly tic a g e n t vrerf em ployed,
3,
P ressure,™
D uring th e f i r s t ex p erim en ts a p r e s s u r e o f 10 l b s ,
p e r square, in c h ..was h e ld , b u t i t was l a t e r n o ted t h a t when th e p re s s u re
was increased..to...l£. l b s , p e r sq u are in c h th e crude f i b e r p e rc e n ta g e was
red u ced c o n s id e ra b ly .
T h e re fo re , most o f th e ex p erim en ts were conducted
a t . a p re s s u re o f.l£ . l b s , p e r sq u are in c h .
The re c o rd e d te m p e ra tu re a t
t h i s p r e s s u r e was l l 60C,
. .Iio..: T im e,-
V arious tim e i n t e r v a l s ra n g in g from £ t o 20 m inutes were
u sed i n t h i s i n v e s t i g a t i o n .
However, i t was found t h a t a tim e i n t e r v a l o f
t e n m in u tes was th e most s a t i s f a c t o r y ,
T h e re fd re , a l l th e ex p erim en ts
re c o rd e d i n t h i s p a p e r e x ce p t th o s e em ploying barium h y d ro x id e were ru n
u s in g a s ta n d a rd tim e in te r v a l, o f TO m in u te s ,
£,
H y d ro ly tic A gents Employed,™
t i c a g e n ts were employed, i n t h i s s tu d y .
V arious acid and a l k a l i n e h y d ro ly ­
The a lk a l in e r e a g e n ts employed
. were sodium -hydroxide, and barium .h y d ro x id e,
S u lf u r ic a c id , hydro c h lo ric
. a c id , and a., m ix tu re of, h y d ro c h lo ric a c id and a c e t i c a c id , were th e a c id
C=,
h y d r o ly tic a g e n ts u s e d .
rJ
0=3
I n a l l c ases th e h y d r o ly tic r e a c tio n s were p re ­
ceded by th e u s u a l e th e r e x t r a c t i o n as d e s ig n a te d i n th e O f f i c i a l M ethods.
The te m p e ra tu re s employed d u rin g th e d ry in g ' and i g n i t i o n o f th e crude
f i b e r r e s id u e s were th e same a s th o s e in d ic a te d i n th e O f f i c i a l M ethods.
6.
G eneral P r o c e d u re .-
Two grams o f th e e th e r e x tr a c te d fe e d m a te r­
i a l were p la c e d ,in a £00 ml E rlenm eyer f l a s k and $0 ml o f th e h y d r o ly tic
a g e n t Were added.
I n a d d itio n , i t was found n e c e s s a ry to add a sm all
amount o f a s b e s to s and a few b o ilin g c h ip s .
The f l a s k s were p la c e d i n th e
a u to c la v e w hich had b een s e t f o r I ^ l b s . p e r sq u are in c h .
The steam was
tu rn e d on and a f t e r th e p r e s s u r e re a ch e d 1$ l b s . th e tim e was n o te d .
The
tim e i n t e r v a l in d ic a te d i n a l l e x p erim en ts was t h a t i n t e r v a l from th e tim e
th e a u to c la v e re a c h e d IJp l b s . p e r sq u are in c h u n t i l i t was tu rn e d o f f .
A f te r c o o lin g , th e f l a s k and c o n te n ts were removed, and th e c o n te n ts f i l ­
t e r e d th ro u g h f i l t e r , c l o t h .
The re s id u e was th e n washed w ith su c c e ss iv e
a p p lic a tio n s , o f w a te r and, f i n a l l y , w ith a c e to n e .
A f te r th e r e s id u e had
been removed from t h e . f i l t e r cloth> i t was d r ie d and i g n i t e d a c c o rd in g
t o th e u s u a l O f f i c i a l M ethods.
I n o rd e r to overcome th e f i l t r a t i o n d i f f i ­
c u l t i e s e n co u n te re d -i n fe e d s Mo. 2 and No. Ii b y f i l t r a t i o n , s e p a r a tio n
by c e n t r i f u g a t i o n was ,u sed .
Tvro grams o f e th e r e x tr a c te d fe e d m a te r ia l
w ere p la ce d , i n . a .100 ml c e n t r i f u g e .tu b e dnd 20 ml o f h y d r o ly tic ag en t
were added to g e th e r w ith some b o ilin g c h ip s and a sm a ll amount o f a sb e s­
to s .
Th'e tu b e was s to p p e re d and p la c e d i n th e a u to c la v e a t th e p re s s u re
and tim e m entioned ab o v e.
The tu b e
Wa1S
removed, co o led , and c e n trifu g e d
f i v e m in u tes = The s u p e r n a ta n t l i q u i d was p o ured o f f and th e re s id u e was
th o ro u g h ly .a g ita te d w ith w a te r and a f t e r c e n tr if u g in g a g a in th e r e s u l t a n t
CO
Q
C=S
S
l i q u i d d e c a n te d o T his p ro c e s s was re p e a te d th r e e tim e s .
in g was w ith a c e to n e ,'
The
f i n a l wash­
The r e s id u e was removed from th e tu b e , d r ie d , and
i g n i t e d a c c o rd in g to th e O f f i c i a l M ethods,
D,
Enzym atic P ro ce d u re Follow ed i n th e C ru d e -F ib e r D e te rm in a tio n ,I n o rd e r to f in d how c lo se tfye chem ical crude f i b e r resem bled th e
d ig e s tiv e crude f i b e r ,, th e enzym atic r e a c t i o n was conducted u sin g p a p a in
and ta k a d i a s t a s e i n a p ro p e rly , b u ff e re d s o lu tio n .
P ro c e d u re ,-
Two grams ■ o f d ry e th p r e x tra c te d , sample were p la c e d i n
a 25>0 ml Eilenm ejrer f la s k and 0,5) grams o f ta k a d ia s ta s e added and 100 ml
o f p h o sp h ate b u f f e r pH lj.,8 .
c u b a to r a t 3J,5>°C,
The m ix tu re was s t i r r e d and p la c e d i n th e i n ­
The fe e d was allo w ed to rem ain i n th e in c u b a to r f o r
jj.8 hours w ith o c c a s io n a l s t i r r i n g .
A f te r 18 h o u rs th e fe e d was removed*
. f i l t e r e d th ro u g h c o a rse f i l t e r p a p e r, and washed w ith d i s t i l l e d w a te r.
The fe e d was re tu r n e d to th e f l a s k and 0,5) grams o f p a p a in w ith a sm all
am ount.of L .c y s tie n e and 100 ml o f p h o sp h ate b u f f e r pH 7 ,2 were added.
The fe e d was k e p t .in th e in c u b a to r 18..h o u rs w ith o c c a s io n a l s t i r r i n g ,
A fte r 18 h o u rs th e fe e d was removed, f i l t e r e d , and washed.
A fte r r e ­
moval. from .the f i l t e r . p a p e r th e r e s id u e was d rie d , and i g n i t e d a c c o rd in g
‘ to th e u s u a l O f f i c i a l M ethods,
IV .
EXPERIMENTAL RESULTS
The r e s u l t s o f th e f i r s t experim ent on fe e d No. I a re show i n Table
II.
The g e n e ra l p ro c e d u re was fo llo w e d u n t i l th e rem oval o f th e f la s k
from th e a u to c la v e when $ ml o f 6 N h y d ro c h lo ric a c id were added to th e
f la s k i n o rd e r to n e u t r a l i z e th e sodium h y d ro x id e rem ain in g i n th e mix­
tu re .
From h e re on th e p ro c e d u re was th e same as d e s c rib e d p r e v io u s ly .
The n o rm a lity o f th e sodium hydro x id e i s th e same as th e n o rm a lity used
by th e O f f i c i a l M ethods.
The in c r e a s e i s a p p ro x im a te ly .05 on each change
o f n o rm a lity .
TABLE I I
Crude F ib e r P e rc e n ta g e s f o r Feed
No. I U sing NaOH as th e H y d ro ly tic
Agent
*
N orm ality
o f NaOH
No. o f
T e sts
Time
M inutes
0 .316
0.316
0.316
0.317
2
2
2
2
10
15
20
5
P re s su re
Ib s /s q .in .
10
10
10
15
Crude F ib e r
P e r Cent
3 Iu 10
W u80
35.00
3U.70
O.M.*
P e r Cent
26.56
2 6 .5 6
26.56
26.56
O f f i c i a l Method
I t w i l l be no ted t h a t th e p e rc e n ta g e crude f i b e r as o b ta in e d h ere was
f a r g r e a te r th a n t h a t o b ta in e d by th e O f f i c i a l M ethods.
The n e x t a n a ly s is was conducted on fe e d No. 3 which resem bled fe e d No.
I i n chem ical c o m p o sitio n .
The g e n e ra l p ro c e d u re was fo llo w e d f o r th e
tim e s in d ic a te d , th e h y d r o ly tic a g e n t and th e p re s s u re were th e
same as
used on fe e d No. 1 6 The r e s u l t s o f t h i s a n a ly s is a re shown i n Table I I I .
— 10 “
TABLE I I I
Crude F ib e r P e rc e n ta g e For Feed
Noe 3 Using NaOH a s th e H y d ro ly tic
Agent
N orm ality
o f NaOH
No. o f
T e sts
Time
M inutes
0.317
0.347
0.347
2
2
2
5
10
5
Crude F ib e r
P e r Cent
P re s su re
Ib s /s q . in .
O.M.
P er Centl
..........
25.00
25.20
28.00
15
15
15
25.51
2 5 .5 1
25.51
The r e s u l t s o f t h i s a n a ly s is show a v e ry c lo se approach th e range o f
th e crude f i b e r p e rc e n ta g e s o b ta in e d by th e O f f i c i a l M ethods,
The r e s u l t s
in d ic a te d t h a t th e tim e o f h y d ro ly s is had l i t t l e e f f e c t on th e r e s u l t s o f
th e a n a l y s i s „
A com plete t e s t o f fe e d No, 3 was conducted u sin g sodium hydroxide o f
th r e e c o n c e n tr a tio n s , and v a ry in g tim e o f h y d r o ly s is .
The r e s u l t s o f
th e s e a n a ly s e s u s in g c o n c e n tra tio n s o f 0 ,3 1 6 N, 0,3li7 N and 0,li07 N sodium
hydroxide and o b ta in e d by tim e s from $ to 20 m inutes show good agreem ent
w ith th e r e s u l t s o f th e O f f i c i a l M ethod,
TABLE IV
Crude F ib e r P e rc e n ta g e s For Feed No, 3 O b tain ed
U sing 1$ l b s , p e r Square In ch P re ssu e and NaOH as th e
H y d ro ly tic A gent_______________________
Time
M inutes
5
10
15
20
Average
No. o f
T e s ts
2
2
2
2
V arying N o rm a litie s
o f NaOH
0 .3 1 6
0.347
0.407
Crude F ib e r P e r Cent
28.10
27.05
28.55
2 9.60
26.70
26.15
26.80
27.40
2 4.90
2 5 .0 0
2 6 .7 0
2 7 .6 0
2 8.30
26.80
26.25
0 . M. Range
•Per Cent
26.12
2 6.15
2 6.15
26.15
"
-
2 5 .5 1
25.00
25.00
25.00
25.00
“
1 1
“
The r e s u l t s shown i n T able IV in d ic a te t h a t th e O f f i c i a l r e s u l t s
could be d u p lic a te d w ith fe e d No. 3 u s in g an e th e r e x tr a c te d sam ple.
Feed No. 2 was a n aly z e d u sin g th e g e n e r a l p ro ced u re and v a ry in g
c o n c e n tra tio n s o f sodium h y d ro x id e .
The r e s u l t s shown i n Table V
in d ic a te t h a t th e lumps formed d u rin g h y d ro ly s is m a te r ia l.
The n o rm a lity
o f sodium hydroxide was l a r g e r th a n p r e v io u s ly in d ic a te d s in c e low er
c o n c e n tra tio n s o f sodium hydroxid e had o b ta in e d no u s a b le r e s u l t s .
TABLE V
Crude F ib e r P e rc e n ta g e s f o r Feeds No. 2
U sing NaOH a s th e H y d ro ly tic A gent, 1$ l b s .
p a r Square In c h P re s s u re and a Time o f 10 M inutes
N orm ality
o f NaOH
No. o f
T e s ts
Crude F ib e r
P er Cent
0 . M.
P er Cent
0 .1 0 7
0 .0 3 0
0.98C
2
2
2
32 .$
2 9 .7
3U.1
7.U3
7.1*3
7.U3
The g e n e ra l p ro c e d u re f o r c e n tr if u g a tio n was fo llo w e d , b u t 10 ml o f
6 N h y d ro c h lo ric a c id was added b e fo re th e c e n tr if u g in g th e f i r s t tim e .
However, th e r e s u l t s as shown i n T able VI in d ic a te d t h a t t h i s method was
not fe a s ib le fo r th is fe e d .
TABLE VI
Crude F ib e r P e rc e n ta g e s f o r Feed No. 2 U sing
C e n tr if u g a tio n f o r S e p a ra tio n , NaOH a s H y d ro ly tic
A gent, 15 l b s . p e r Square In ch P re s s u re and a Time o f
10 M inutes
N orm ality
o f NaOH
0.1*07
0.615
0 .830
No. o f
T e sts
3
3
3
Crude F ib e r
P e r Cent
0 . Me
P e r Cent
39.95
1*1.65
39.50
7.1*3
7.1*3
7.1*3
“ ■
12
“
The r e s u l t s o f th e sodium h y d ro x id e a n a l y s i s o f f e e d s No. I and No. 3
i n d i c a t e d t h a t th e a l k a l i h y d r o ly s is w orked w e ll f o r f e e d s w ith h ig h cru d e
f i b e r p e r c e n ta g e s , b u t t h e r e s u l t s o f f e e d s No. 2 and No. it i n d i c a t e d t h a t
sodium h y d ro x id e was n o t th e p r o p e r h y d r o l y t i c a g e n t .
A s e r i e s o f e x p e rim e n ts w ere p e rfo rm e d u s in g b ariu m h y d ro x id e a s th e
h y d ro ly tic a g e n t.
B arium h y d ro x id e s o l u t i o n s w ere p r e p a r e d h a v in g n o rm al­
i t i e s o f 0 .1 6 0 and 0 .3 2 0 .
F eed s No. I and No. 2 b e in g t y p i c a l exam ples
o f ea c h g rou p w ere h y d ro ly z e d w ith b ariu m h y d ro x id e a t I 5 l b s . p e r s q u a re
in c h p r e s s u r e f o r tw e n ty m in u te s .
The r e s u l t s a s shown i n T ab le V II i n ­
d i c a t e d a d e c r e a s e i n o r g a n ic r e s id u e when t h e h ig h e r c o n c e n tr a ti o n o f
b ariu m h y d ro x id e was u s e d .
TABLE V II
Crude F ib e r P e rc e n ta g e s O b ta in ed Using
B a(0H)2 as th e H ydrolyzing A gent, 15 l b s .
I)er Square In ch P re s s u re and a Time o f 20 M inutes
Feed
No. o f
T e s ts
I
I
2
2
2
2
2
2
N orm ality
Ba(OH) 2
0 .1 6 0
0 .32 0
0.160
0.320
Crude F ib e r
P er Cent
0 . M0
P er Cent
U .00
39,16
32.15
28,70
26.56
26.56
7.4 3
7.43
The e x p e rim e n ts u s in g b a riu m h y d ro x id e 0 .3 £ ? N was co n d u c te d on f e e d s
Nb. I and No. 2 and t h e r e s u l t s showed f u r t h e r r e d u c t io n i n t h e
f i b e r p e r c e n ta g e .
The r e s u l t s a r e shown i n T a b le V I I I .
cru d e
T 19 TABLE V XIl
Crude F ib e r P e rc e n ta g e s U sing 0<,35B N
Ba(OH) 2 a s H y d ro ly tic A gent51 19 l b s . p e r
' Square In c h P r e s s u r e ■and a Time o f 30 M inutes
.1
2.
%
i
.
Crude- F ib e r
Per. .Cent. .
T e s ts
F eed
3lu35
. 2H.20 .
.2
2 ■.
..
Oo Mo
P e r C ent
!
26.56
7oh3
Ho f u r t h e r ex p erim en ts w ere conducted u sin g barium h y d ro x id e as th e
n o rm a lity o f th e l a s t s o lu tio n was 0 o3!?£ N and no h ig h e r n o r m a litie s co u ld
be o b ta in e d u s in g Ba(OH)^0SHgO u n le s s a warm s o lu tio n was u s e d .
HAND­
BOOK OF CHEMISTRY AND PHYSICS (10) i n d i c a t e s t h a t 5 oh .g o f Ba(OH)2 0SHgO
d is s o lv e s i n 100 ml o f w a te r a t lB °C .
As room te m p e ra tu re i s g e n e r a lly
ab o u t 2h° th e s a t u r a t i o n p o in t had ab o u t b e en re a c h e d .
The n e x t group o f h y d r o ly tic a g e n ts u sed was th e a c i d s .
The f i r s t
a c id employed was th e 0.2BB N s u l f u r i c a c id o f th e O f f i c i a l M ethods.
in c r e a s e i s a p p ro x im a te ly .05 on each change o f n o rm a lity .
The
The fe e d s
t e s t e d w ere Ho. 2 and No. L as fe e d s No. I , a n d .No. 3' h a d -b e e n t e s t e d
s u c c e s s f u lly w ith..sodium h y d ro x id e . ..The g e n e r a l p ro c e d u re was fo llo w ed
u s in g s u l f u r i c a c id , 0 .2 5 5
f o r v a ry in g tim e s a t 15 ID s. p e r sq u are
in c h p r e s s u r e .
The e x p e rim e n ts w ere c o n tin u e d u s i n g H ig h e r . c o n c e n tr a ti o n s o f s u l ­
f u r i c a ,c id .
T a b le IX and
The r e s u l t s o f th e s u l f u r i c a c i d e x p e rim e n ts a r e shown i n
g iv e p e r c e n ta g e s o f a lm o s t d o u b le th o s e o b ta in e d b y th e
O f f i c i a l M eth o d s.
-■
•
/
TABLE IX
Crude F ib e r P e rc e n ta g e s O b tain ed Using
S u lf u r ic Acid as H y d ro ly tic Agent and l5
l b s , p s r Square In c h P re s su re
Feed
No. o f
T e sts
N orm ality
H2SOli
Time
M inutes
Crude F ib e r
P er Cent
2
2
2
2
h
2
k
2
2
2
2
2
2
2
0 .2 5 5
0 .255
0 .255
0 .3 0 3
0 .303
0.352
0.352
5
10
15
5
5
5
5
13.95
12.45
11.30
14.00
7.7 0
14.40
7.3 5
. 0 . M.
P er Cent
7.43
7.43
7.43
7.43
3 .5 6
7.43
3 .5 6
Feed No, L was a n aly z e d u sin g s u l f u r i c a c id o f th r e e c o n c e n tra tio n s
and v a ry in g tim e s a t 1$ l b s , p e r sq u are in c h p r e s s u r e .
shown i n Table X and in d i c a t e no r e d u c tio n i n th e
The r e s u l t s a re
crude f i b e r p e rc e n ta g e
was accom plished by e i t h e r in c r e a s in g th e c o n c e n tra tio n o f th e h y d ro ly tic
a g en t o r th e tim e o f th e
re a c tio n .
TABLE X
Crude F ib e r P e rc e n ta g e f o r Feed No, k
U sing S u lf u r ic Acid a s th e H y d ro ly tic Agent
and 15 l b s , p e r Square In c h P re s s u re
Time
M in u tes
No. o f
T e s ts
5
10
15
20
2
2
2
2
A verage
V ary in g N o r m a litie s
o f H2SO,
0 .255
0 .3 0 3
0.352
Crude F ib e r P e r Cent
7 .1 0
6 .9 0
6 .8 5
7.35
6 .5 0
6.4 5
7.35
6 .9 0
6 .4 0
7.1 0
6 .6 5
6 .5 0
7.05
6 .9 0
6 .6 5
0 . M. Range
P e r C ent
3 .8 5
3 .8 5
3 .8 5
3 .8 5
-
3.0 5
3.0 5
3.o5
3.05
3.5 6
As th e r e was no g r e a t re d u c tio n o f th e av erag e p e rc e n ta g e to th e
O f f i c i a l p e rc e n ta g e th e e x p erim en ts u s in g s u l f u r i c a c id were n o t c a r r ie d
fu rth e ro
H y d ro c h lo ric a c id a lo n e and i n com bination w ith a c e t i c a c id was used
as th e h y d r o ly tic a g e n t.
The g e n e ra l p ro c e d u re was fo llo w ed u s in g th e
fo llo w in g a c id r e a g e n ts :
(A) a m ix tu re o f
ml 0 .2 6 6 N h y d ro c h lo ric a c id
and 50 ml 0 .2 9 0 N a c e t i c a c id ; (B) 0.b97 N h y d ro c h lo ric a c id and (C) 0.976 N
h y d ro c h lo ric a c id .
The tim e f o r th e h y d ro ly s is was 10 m in u te s and th e
p r e s s u r e 1$ l b s . p e r sq u are in c h .
The r e s u l t s a re shown i n Table XI.
TABLE XI
Crude F ib e r P e rc e n ta g e s U sing Acid R eagents
Feed
I
2
3
h
No. o f
T e sts
2
2
2
2
REAGENTS
C
A
B
Crude F ib e r Pei Cent
3 3.30
13.35
37.70
7 .9 0
3 2.75
13.25
33.15
7.2 5
32.80
12.70
32 .UO
7 .5 0
0 . Me
P e r Cent
26.56
7.U3
2 5 .5 1
3 .5 6
The r e s u l t s o f th e s e e x p erim en ts gave cru d e f i b e r p e rc e n ta g e s con­
s id e r a b ly above th e O f f i c i a l Method p e rc e n ta g e s and th e s e ex p erim en ts
were n o t c o n tin u e d .
A fte r th e com pletion o f th e a c id a n a ly s is o f th e f e e d s , a combina­
tio n
o f a c id and a l k a l i re a g en s was u s e d .
The f i r s t a n a ly s is o f t h i s
ty p e was conducted u s in g th e re a g e n ts o f th e O f f i c i a l M ethod, b u t in s te a d
o f b o i l i n g th e fe e d m a te r ia l f o r t h i r t y m in u tes th e fe e d s were h y d ro ly zed
a t 15> l b s . p e r sq u are in c h p r e s s u r e f o r t e n m in u te s .
The fe e d m a te r ia ls
16 ■=
were washed and f i l t e r e d betw een a c id and a l k a l i h y d r o ly s is o The crude
f i b e r p e rc e n ta g e s o b ta in e d were s l i g h t l y below th e O f f i c i a l Method p e r­
c en ta g e s i n th e case o f fe e d s No= I , No, 2, and No, 3, w h ile fe e d No, I*
was w ith in th e O f f i c i a l ran g e as t h i s p e rc e n ta g e approached th e h ig h e s t
O f f i c i a l Method p e rc e n ta g e .
A nother a n a ly s is was
conducted u s in g s tro n g e r
re a g e n ts and th e crude f i b e r p e r c e n t f o r fe e d No, ii went down.
Table
XII shows th e crude f i b e r p e rc e n ta g e o b ta in e d from th e s e a n a ly s e s and th e
v a r i a t i o n s from th e O f f i c i a l Method p e rc e n ta g e ,
TABLE H I
Crude F ib e r P e rc e n ta g e s U sing I 5 l b s , p e r
Square In c h P re s s u re f o r 10 M inutes D uring
Both Acid and A lk a li H ydrolyzing P e rio d s
Feed
No, o f
T e s ts
I
2
2
2
2
2
2
2
3
2
U
2
I
2
3
U
Crude F ib e r
P er Cent
R eagents
R eagents o f 0 ,
R eagents o f 0 ,
R eagents o f 0 ,
R eagents o f 0 ,
0 .2 5 N H2SOj
O.iiO N NaOHa
0 .5 0 N HCl
0.U0 N NaOH
0 .5 0 N HCl
0 .3 2 N NaOH
0 .5 0 N HCl
0.U0 N NaOH
M,
M,
M,
M,
V a ria tio n s From
0 . M. Average
20.65
5 .7 0
19 o75
5 .1 0
-5 .9 1
-1 .7 3
-5 .7 6
+ 1 .5 1
21.55
-5 .0 1
5 .6 0
-1 .8 3
18.75
-6 .7 6
3.5 5
- 0 .0 1
I t i s e v id e n t from th e r e s u l t s o f t h i s l a s t s e r i e s o f ex p erim en ts
t h a t t h i s method o f a n a ly s is i s th e b e s t o f th e
methods u sed f o r a l l o f
th e fe e d m a te r ia ls i n o b ta in in g p e rc e n ta g e s n ear th e ran g e o f th e O f f i c i a l
p e rc e n ta g e s .
W ith th e c o n c lu s io n of* th e p r e c e d in g e x p e rim e n ts i t was th o u g h t ad­
v i s a b l e t o check t h e cru d e f i b e r p e r c e n ta g e s b y u s in g enzym es.
p e rim e n ts w ere c o n d u cted on d u p l i c a t e s a m p le s .
The ex­
T h is a n a l y s i s f o llo w e d th e
p ro c e d u re g iv e n f o r e n z y m a tic a n a l y s i s i n e x p e r im e n ta l p r o c e d u r e .
The
r e s u l t s o f t h i s a n a l y s i s i n T ab le X III gave th e cru d e f i b e r p e r c e n ta g e s
f a r g r e a t e r th a n th o s e o b ta in e d by th e O f f i c i a l M ethod.
TABLE X III
Crude F ib e r P e r c e n ta g e s U sing E nzym atic A gents
Feed
I
2
3
U
Crude F ib e r
P e r Cent
0 . M.
P e r C ent
V a r i a t i o n s From
O.M. P e r Cent
$3 .ItO
31.30
51.70
2 0 .6 0
26.56
7.L3
2 5 .5 1
3 .5 6
+ 26.8b
+23.87
+26.19
+ 17.0b
I t i s no ted from th e t a b l e t h a t fe e d s No. I and No. 3 have a crude
f i b e r p e rc e n ta g e o f double th e O f f i c i a l p e rc e n ta g e , and fe e d s No. 2 and
No. b have a p e rc e n ta g e o f ab o u t fo u r tim e s th e O f f i c i a l p e rc e n ta g e s .
- 18 Vo
DISCUSSION
I n t h i s d i s c u s s i o n t h e m ethod o f t h e O f f i c i a l cru d e f i b e r d e te rm in e ■
a t i o n w i l l b e com pared w ith th e m ethod em ployed in , t h i s i n v e s t i g a t i o n s '
-The m echanism s o f th e r e a c t i o n s w i l l a l s o b e d is c u s s e d and th e -u se o f
v a r io u s r e a g e n ts em ployed i n t h e cru d e f i b e r d e t e r m in a tio n .
The tim e o f t h e h y d r o ly s is a s g iv e n i n t h e o f f i c i a l m ethods i s t h i r t y
m in u te s f o r . b o th a c i d an d a l k a l i r e a c t i o n s 0 -■ Though th e o f f i c i a l 'm e t h o d o f
a n a l y s i s i s u s e d f o r th e cru d e f i b e r d e t e r m in a tio n i n . M orttanay no way o f
c o m p en sa tin g f o r d i f f e r e n c e s - i n 'a t m o s p h e r i c p r e s s u r e i s ta k e n in t o con­
s id e ra tio n s
As t h e te m p e r a tu r e o f th e b o i l i n g p o in t o f a s o l u t i o n goes
down ,a p p ro x im a te ly 6 ° C p e r k il o m e t e r r i s e . i n a l t i t u d e , i t i s e v id e n t t h a t
b o i l i n g .te m p e r a tu r e o f t h e h y d r o ly t ic , a g e n ts w i l l b e l e s s i n M ontana th h n
i n th e c o a s ta l r e g io n s o
B id w e ll ( l ) p o in t e d o u t m arked v a r i a t i o n s a n th e
d i f f e r e n t l a b o r a t o r i e s th ro u g h o u t t h e c o u n try and F ra p e s (7 ) r e p o r te d
v a r i a t i o n s i n t h e c ru d e f i b e r r e s u l t s b y l a b o r a t o r y lo c a tio n s *
Though
t h e s e r e p o r t s w ere made m ore th a n 2$ y e a r s a g o .n o th in g was done i n o r d e r
t o s ta n d a r d i z e t h e 't e m p e r a t u r e u s e d i n th e cru d e f i b e r d e te r m in a tio n * >
From a l o g r i t h m e t i c r e p r e s e n t a t i o n o f th e A rrh e n iu s e q u a tio n ,
lo g b
=. lo g
A
”
t h e e f f e c t o f te m p e r a tu r e ch an g es on t h e r e a c t i o n r a t e can r e a d i l y be
s e e n f o r As E / and E a r e c o n s ta n ts and o n ly T v a r i e s ,
T h e re fo re ^ an y
change i n t h e te m p e r a tu r e o f th e r e a c t i o n ch an g es th e r e a c t i o n r a t e i n
d i r e c t r e l a t i o n , i c e * , i n c r e a s e i n te m p e r a tu r e in c r e a s e s t h e r a t e and
.d e c re a s e i n te m p e r a tu r e d e c r e a s e s th e r a t e *
F o r t h i s r e a s o n t h e a u to c la v e
i
“ 19 =
was u se d i n o r d e r t o i n c r e a s e th e te m p e r a tu r e o f t h e r e a c t i o n d u r in g t h e
h y d r o ly tic p ro c e ss*
The te m p e r a tu r e , o f t h e r e a c tio n , a s r e c o r d e d b y th e
a u t o c l a v e . th e rm o m eter was I l d 0Co
T h is i n c r e a s e i n p r e s s u r e n o t o n ly i n c r e a s e s .th e b o i l i n g p o in t o f
t h e m ix tu r e , b u t d e c r e a s e s t h e tim e o f h y d r o ly s is a t t h e maximum te m p e r/
a tu re „
The volum e o f t h e h y d r o l y t i c a g e n t u se d b y t h e O f f i c i a l M ethod
i s 200 ml f o r . b o t h a c i d and a l k a l i *
The volum e, u s in g a n a u to c la v e p r e s s u r e
o f 10 t o 19 l b s * p e r sq u are! in c h , i s £>0 m l, a d e c r e a s e o f 150 m l o f
■«.
' v
r e a g e n t » The u se- o f a s m a lle r volum e o f " a c id and a l k a l i i n t h e in v e s ­
t i g a t i o n c a u s e s no l a r g e d e v i a t i o n from t h e l a r g e volume o f t h e O f f i c i a l
M ethods n o t u s in g p r e s s u r e ,
A f a c t o r c o n s id e r e d h e re i n t h e u s in g o f j?0
ml o f r e a g e n t was t h e .b o ili n g o v e r o f th e m ix tu r e w h ile t h e p r e s s u r e was
d e c r e a s in g ,.
The tim e ; o f h y d r o l y s i s o f t h e O f f i c i a l Method, i s 30 m in u tes' w ith t h e
m ix tu re b o i l i n g , f o r t h a t .tim e .or, .a t .a maximum te m p e r a tu r e f o r 30 m in u te s ,
By using, a n a u to c la v e i n w h ich to . co n d u ct t h e h y d r o ly s is t h e tim e o f
h y d r o ly s is i s re d u c e d t o 10 m in u te s ,
From th e e x p e r im e n ta l r e s u l t s th e
changes i n tim e , w ere shown t o have no e f f e c t on th e am ount o f cru d e f i b e r
o b ta in e d from e a c h f e e d .
However, a tim e o f 10 m in u te s a t maximum temp­
e r a t u r e was s e l e c t e d a s m o st s a t i s f a c t o r y f o r a l l f e e d s ,
W intoh (1 9 ) b e g in s h i s cru d e f i b e r a n a l y s i s b y s a y in g **E x t r a c t a 2
gram sam ple w ith ether** and t h i s c o n tin u e s w ith th e u s u a l p r o c e d u r e ,
a g r i c u l t u r a l c h e m is tr y t e x t o f t h e same d a t e b e g in s th e d i s c u s s i o n o f
cru d e f i b e r a n a l y s i s i n th e same w ay th o u g h h y d r o c h lo r ic a c i d i s u sed
i n s t e a d o f s u l f u r i c a c id i n t h e p r o c e d u r e ,
B id w e ll and H dpdt ( 3 ) ,
Ah
'
=
20
==
use e th e r e x t r a c t i o n to p re c e d e th e cru d e f i b e r d e te r m in a tio n .
I n th e
methods u s in g in o rg a n ic a c id s and a l k a l i re a g e n ts f b r th e m o si p a r t the.
e th e r e x tra c tio n -w a s used*
The c h ie f re a s o n f o r th e rem oval o f th e f a t
m a te r ia l by e th e r e x t r a c t i o n i s t h a t i f th e f a t s a re n o t removed th e '
fo rm a tio n o f soaps w ith th e a l k a l i re a g e n t make, s e p a r a tio n o f th e re s id u e
much, more d i f f i c u l t .
T his i s e s p e c i a l l y t r u e i f . .a ..h ea v ie r a l k a l i i s used
a s th e .h y d r o ly tic , a g e n t ,,. A lso,, t h e r e a re ..few. fe e d s, .which w i l l
be an aly zed
f o r crude f i b e r o n ly .
Changes i n th e c o n c e n tra tio n o f re a g e n ts have been' su g g e ste d b y J',
H0 and E , Vi, V oelcher (1 8 ), 'W, L ep p e r . (13). and o t h e r s ," The c o n c e n tra tio n s
have v a lu e s from 2 ,0 p e r c e n t t o 3 *12£ p e r c e n t f o r b o th a c id and a l k a l i
r e a g e n ts .
The r e s u l t s o f th e s e ex p erim en ts u sin g a h ig h e r c o n c e n tra tio n
. in d ic a te s t h a t th e tim e o f th e r e a c t i o n may be red u ced as much as te n
m in u te s .
A lso th e volume o f th e re a g e n ts may be red u ced u s in g .a h ig h e r
c o n c e n tra tio n o f a c id and a l k a l i .
j.
From th e e x p e rim e n ta l r e s u l t s i t has '
,
b e en shown t h a t b o th a c id and a l k a l i h y d r o ly s is i s n e c e s s a ry i n o rd e r to .
d u p lic a te th e o f f i c i a l r e s u l t s on a l l ty p e s o f fe e d sam p les. I t .has been
,'
, shown tt h a t a c id removes some, o f th e non fib r o u s , m a te r ia l b u t no t a l l o f
it,
vC/''
At th e same tim e th e a l k a l i removes most o f th e non f ib r o u s m a te r ia l
i n th e hay ty p e fe e d s , b u t r e m o v e s . l i t t l e o f th e same m a te r ia l i n th e case
o f g fd in and mixed sam p les.
The a c id h y d ro ly s is o f c a rb o h y d ra te s re d u c e s th e s ta r c h e s and su g ars
to sim ple sugahs i n m ost c a se s g lu c o s e . A cids i n g e n e ra l do n o t . e f f e c t
.
■
'
■
'
th e c e ll u lo s e m olecule u n le s s th e h y d ro ly s is i s conducted o v e r a long
tim e i n t e r v a l i n w hich c a s e s some f r a g m e n ta tio n o c c u r s .
a
A l k a l i a g e n ts
■=
21
■=
r e a c tin g w ith th e s ta r c h m olecule cause th e fo rm a tio n o f a g e l .
form ed i s not f i l t e r a b l e „
The■g e l
The u se o f a l k a l i i n h y d ro ly z in g fe e d s No6 2'
and No6 L caused th e fo rm a tio n o f lumps i n th e m ix tu re which rem ained a t
th e end of. th e ' h y d ro ly z in g p e r io d ,
Tiihen th e lumps .were b ro k en open w ith
a s t i r r i n g ro d th e r e was no eviden ce o f a r e a c tio n h aving ta k e n p la c e 6
A lk a li re a g e n ts have l i t t l e e f f e c t on th e c e llu lo s e m o lecu les e x ce p t o v er
lo n g tim e i n t e r v a l s , i n which ..case fra g m e n ta tio n o c c u r s ,... However, th e
c o n c e n tra tio n , o f th e a l k a l i m ust be g r e a te r th a n th e ones u sed i n t h i s
work c
P r o te in s .are r e a d i l y h y d ro ly zed i n t o . . sim ple, amino a c id s and s o lu b le
.p eptides, by b o th , s u l f u r i c and h y d ro c h lo ric a c id .
For q u a n t i t a t i v e hy­
d r o l y s i s , tim e s o f s e v e r a l h o u rs a re u sed to b o i l th e a c id s c o n ta in in g
th e p r o t e i n .
The r e a c tio n o f th e a c id w ith th e p r o t e i n m o lecu le p ro b a b ly
caused in co m p leted h y d ro ly s is to amino a c id s d ip e p tid e s , p e p to n e s , p o ly ­
p e p tid e s , e t c , , s o lu b le i n th e wash w ater.,
P r o te in s a re r e a d i l y h y d ro ly zed b y a l k a l i r e a g e n ts ,
The a l k a l i
h y d ro ly s is i s v e ry r a p id and w i l l work r e a d i l y i n cases as crude f i b e r
a n a ly s is where th e q u a l i t a t i v e .and q u a n t i t a t i v e a n a ly s is o f p r o te in s i s
n o t d e s ir e d ,
The enzym atic a n a ly s is o f th e sample was conducted i n o r d e r to see
how c lo s e to a b io lo g ic a l, crude, f i b e r th e a c id and a l k a l i h y d r o ly tic
. end p ro d u c ts w ere a p p ro a c h in g .
The. m ain o b je c tio n to th e enzym atic
a n a ly s is , i s . i t s slow ness as compared w ith in o rg a n ic .m e th o d s i,
Woodson
and M ackenzie (.20) used th e enzyme p a n g e s tin to d eterm in e crude f i b e r
p e r c e n ta g e s .
They found t h a t th e v a lu e s o b ta in e d f o r th e crude f i b e r
— 22 =
■p e r c e n ta g e s w ere a b o u t f o u r tim e s th e amount o b ta in e d w ith t h e o f f i c i a l
m e th o d so
The r e s u l t s o f my p a p e r w ere W ell i n k e e p in g w ith t h e r e s u l t s
o b ta in e d i n t h i s p a p e r c
The e t h e r e x t r a c t e d f e e d was f i r s t t r e a t e d w ith T a k a - d ia s ta s e to
-"remove th e c a r b o h y d r a te m a t e r i a l ,
-A fte r t h e f e e d had beeri w ashed p a p a in
was ad d ed to th e sam ple w ith a sm a ll, .amount o f c y s te in e t o a c t i v a t e th e
enzym e.
The p a p a in rem oves t h e p r o t e i n m a t e r i a l .
A f te r t h e s e r e a c t i o n s
have b e e n c o m p lete d t h e cru d e f i b e r i s o b ta in e d i n t h e u s u a l w ay.
A f te r com paring t h e o f f i c i a l m ethods w ith th e e n z y m a tic m ethod, i t
i s s e e n t h a t th e
o r g a n ic re s id u e :, c a l l e d cru d e f i b e r , i s n o t t h e same
i n b o th .m e th o d s .
The e n z y m a tic m ethod rem oves c a r b o h y d r a te s and p r o t e i n s
w h ile th e a c i d an d . a l k a l i r e a g e n ts rem ove t h e s e p r o d u c ts and
o th e r m a te ria ls
p o s s ib ly
= 23 "
V I,
SUMMARY
From th e e x p e rim e n ta l r e s u l t s o f t h i s s e r i e s o f e x p erim en ts th e
fo llo w in g c o n c lu s io n s can be s ta t e d :
1.
Crude f i b e r i s c o n sid e re d a v a r ia b le q u a n tity dep en d en t on th e
J
f o llow ifig f a c t o r s :
(a ) th e te m p e ra tu re o f th e r e a c tio n m ix tu re , (b) th e
c o n c e n tra tio n o f th e re a g e n t o r re a g e n ts u sed , and (c ) th e ty p e o f re a g e n t
u se d .
2 . ,,R e s u lts ,o f th e O f f i c i a l Method on fe e d s w ith a. h ig h crude f i b e r
c o n te n t may be approxim ated by th e u se o f a s in g le h y d r o ly tic re a g e n t a t
e le v a te d te m p e ra tu re s .
'3 .
I n fe e d s c o n ta in in g l a r g e r p r o p o r tio n s o f s t a r c h b u t low crude
f i b e r p e rc e n ta g e s a c c o rd in g to th e t i W i c l & l 'fifefc'Hods, u n r e lia b le r e s u l t s
a re 'a b ta ih e d ’u s ih g .a s in g le re a g e n t a t e le v a te d ' te m p e ra tu re s , and
; Ifc
Enzym atic h y d r o ly s is of. fe e d m a te r ia ls em ploying th e com bination
o f p r o t e o l y t i c and c a rb o h y d ra te s p l i t t i n g enzymes were found to y ie ld a
, c o n s id e ra b ly h ig h e r q u a n tity o f o rg a n ic re s id u e s , th a n .o b ta in e d b y th e
chem ical means, o f th e O f f i c i a l M ethods.
.• :
■•
;'
•• ■ ' V:
V IIb
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Investigations on the deterfflination of crude fib e r
date
I S S U E D TC
I ___
L
j~
••••
/V375
m
s s s
--,
O O yP - 2_