SEPTEMBER~
1964
HITZMAN:
595
R A P I D 1V[EASUI~EMENT OF D E T E R G E N T B I O D E G R A D A B I L I T Y
idly and that adequate numbers of microorganisms are present in all samples to cause biodegradability. The initial inoeulum is not critical as long as it is adequate to cause the lauryl sulfate to degrade in two days. Increasing the inoeulum beyond this point does not seem to cause an appreciable increase in the rate of degradation and it does not cause a more complete degradation. Final levels of degradation are the same as in the standard river die-away test, with the ae-
celerated test simply compressing the test time to less than one-half. REFERNbrCNN 1, I-Iammerton, G., J. Appl. Chem. 6, 517 (1955). 2. Sawyer, G. N., t~. It. Bogan and J. t~. Simpson, Ind. Eng. Chem. 48, 2, 236 (1956). 3. The Procter & Gamble Co., personal communication. 4. Standard ~Iethods for Examination of W a t e r a n d Wastewater, l l t h :Editio;~, P a r t 1, Surfactants (Anionic) 1960, p. 246-251. [:Received M a r c h
13, 1 9 6 4 - - - A c c e p t e d
May
8, 1 9 6 4 ]
Determination of the Glyceride Structure of Fats; Analysis of 14 Animal and Vegetable Fats M. R. SUBBARAM 2 and C. G. YOUNGS, National Research Council of Canada, Prairie Regional Laboratory, Saskatoon, Saskatchewan, Canada Abstract The glyceride compositions of seven animals and seven vegetable fats have been determined by GLC analysis of the oxidized esterified glycerides as described in an earlier paper in this series. The compositions determined are compared with those calculated from lipase hydrolysis data according to the method of VanderWal. Good agreement was found between the calculated and determined compositions for the majority of the 14 fats. The exceptions were human fat and the more saturated vegetable fats, pahn oil and cocoa butter, where some discrepancies occurred.
Introduction T
tiE OZ~CaNabYSIS Of oxidized esterified glycerides (1) provides a rapid and accurate means of determining the f a t t y acid distribution in natural fats. Initial investigations on a few fats showed good agreement between the determined glyceride composition and that calculated from lipase hydrolysis data according to the method of VanderWal (2). The present work extends this type of analysis to 14 animal and vegetable fats. The results obtained are compared with those calculated from lipase hydrolysis data.
Experimental The vegetable oils studied were commercial, refined and bleached samples. The anhnal fats were coldextracted with chloroform from fresh adipose tissue. Twenty mg of each fat was oxidized with permanganate-periodate as described earlier (1). The resulting azelao-glycerides were methylated with diazomethane and the oxidized esterified glyeerides analysed by GLC. Conditions for GLC analysis and methods of calculation were as previously described (1). The original f a t t y acid compositions of the fats were determined by GLC of their methyl esters on an o-phthMie-ethylene glycol column. The method of Youngs (3) was used for lipase hydrolysis in which the composition of the liberated f a t t y acids was determined rather than that of the unhydrolysed monoglyeerides. All results are reported as mole percentages.
Results and Discussion Tables I and I I give the glyeeride compositions as determined and as calculated from lipase hydrolysis data. Since the separation of the oxidized esterified Issued as NRC No. 8052. s National Research Council of Canards Postdoctorate Fellow.
glyeerides by GLC is dependent on their effective carbon nmnber, those glyeerides giving rise to the same carbon number emerge together and are determined as a group as indicated in the tables. The individual glycerides can be calculated from lipase hydrolysis data and this has been done. Comparisons are then made on the basis of the sum of the calculated glyeerides in a group where two or more glyeerides have the same carbon nmnber. Since myristie acid is a minor component in the fats investigated, the calculated proportion of myristic-eontaining glycerides is small. Glycerides containing more than one myristic acid are less than 0.1%. Tables I i I and IV give the original f a t t y acid compositions of the fats, as determined by GLC of their methyl esters, and those calculated from the glyceride compositions obtained. The agreement between these compositions for the individual fats serves as a check on the glyceride analysis. The f a t t y acid composition in the 1-3 positions is also given in the Tables. These latter figures were used in calculating the glycride cmnpositions on the basis of VanderWal's theory. Good agreement between the determined and calculated compositions was found for linseed, corn, olive, cottonseed and soybean oils. F o r the more saturated fats, cocoa butter and palm oil, the proportion of disaturated glycerides found was somewhat higher than that calculated, with the remaining glycerides being lower than calculated. In view of the good agreement obtained for the other vegetable fats, this suggests that the actual glyeeride distribution for the more saturated vegetable fats may be slightly different than that predicted from lipasc hydrolysis data. This however, requires f u r t h e r investigation. Agreement between the calculated and determined glyeeride compositions for the animal fats was generally good with the exception of human fat. In the latter ease considerably more monosaturated glycerides were found than would be expected from lipase hydrolysis data, with a corresponding drop in the proportions of the fully unsaturated glycerides and more saturated glycerides. A similar pattern was found for two other samples of human fat. Since humans undoubtedly receive a much higher proportion of dietary fat than the other animals tested, this discrepancy may represent the effect of combined endogenous and exogenous fats. In general it appears that glyceride composition calculated on the basis of lipase hydrolysis data provides a good estimate for the majority of natural
596
THE
JOURNAL
OF
THE
OIyceride Carbon
No.
AMERICAN TABLE Composition
. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
U--unsaturated
acids;
M--myristic
acid;
No.
U--unsaturated
Fatty
acids;
M--myristic
42
44
46
48
50
52
54
Us
MU2
PUs MuU
SU~ MPU Ms
P2U MSU M2P
PSU MPe MeS
SU~ P3 MPS
P~S MS2
S~P
Ss
5.9 9.4
0.9 1.1
30.5 29.9
7.6 6.1
35.6 30.1
8.8 9.1
9.1 9.8
1.3 3.8
0.3 0.5
.... ,..
73.0 69.6
.... .
17.3 18.9
8.0 8.4
1.0 1.5
0.6 1.3
6.3
60.5 60.2
.
30.0 31.1
4.3 3.8
4.0 4.4
1.1 0.3
0.2 0.3
.... 0.1
6.2 9.7
8.8 11.3
19.1 15.1
39.9 35.3
22.7 21.3
1.4 2.2
0.8 1.6
..
59.6
Palm oil Original .......................... Calc ..................................... 1-3 positions .................. Linseed oil Original ............................ C a l c ................................. . . . 1 - 3 p o s i t i o n s ..................... Corn oil Original ........................... Calc .................................. 1-3 positions .................... Cocoa butter O r i g i n a l ........................... . . . Calc .................................... 1-3 positions .................... Olive oil Original ............................. Calc .................................... 1-3 positions ................... Cottonseed oil O r i g i n a l .......................... . . . C a l c ................................. . . 1-3 positions .................... Soybean oil Original ............................. Calc .................................. 1-3 positions acids;
P
_.
28.7
6.0
1.4
O~Z
27.1 26.6
2.0 1.5
42.4 43.6
6.5 5.8
18.0 18.8
3.0 3.0
1.O 0,7
. . . . . . . . . .
55.5 56.5
....
28.0 27.2
9.2 10.0
4.8 3.3
2.3 2.4
0.2 0.6
. . . . . . . . . . . .
and
S--stearic
. . . . . . . . . . . . . . . . . . . . . . .
.
.
.
.
.
.
.
.
.
.
.
.
acid.
TABLE II C o m p o s i t i o n of A n i m a l
Fats
36
38
40
42
44
46
48
50
52
54
U3
MU2
PUs MsU
SUs MPU Ms
P~U MSU M~P
PSU MPe M~-S
SU~ Pa MPS
P~S MS2
SeP
Ss
21.8 32,0
3.6 3.4
45.7 34.5
13.6 10.1
12,6 11.9
2.7 5.4
ii~
0.7
O:i
....
4.4 4.2
0.9 2.0
0.4 0.8
.... ....
20.3 22.8
4.2 4.4
24.8 25.8
20.7 17.1
14.1 11.7
10.2 10.8
55.0 55.8
3.0 2.0
33.1 31.6
4.2 4.0
4.2 5.2
0.5 1.1
o:g
31.6 31.7
1.5 1.6
36.7 34.9
10.6 11.1
12.9 11.9
4.7 6.3
1.6 1.7
0.4 0.7
0.1
.
1.8 2.2
2.2 2.4
01~
::::
:iii
....
.
-
14.5 13.6
2.3 2.0
38.0 37.1
14.3 12.7
6.7 7.7
18.3 19.7
1.9 2.4
27.6 31.1
3.6 4,3
36.4 34.2
13.5 9.5
12.0 13.4
4.3 4.9
2.0 2.0
0.6 0.6
........ ........
31.1 33.0
.... ....
34.3 35.9
14.0 9.2
13.2 13.7
5.0 6.0
2.0 1.3
0.4 0.9
. . . . . . .
P--pahnitic
acid
and
S--stearic
acid. Fatty
Fats S 6.3 5.7 6.5
46.5 45.9 39.4
.... .... ....
7.8 6.8 10.2
3.4 2.9 5.1
88.8 90.3 84.7
.... .... ....
13.5 13.1 17.7
1.8 1.9 2.5
84.7 85.0 79.8
.... .... ....
28.1 29.1 40.1
32.9 32.8 46.8
38.3 38.1 13.1
.... .... ....
12.6 12.7 19.0
2.6 2.3 4.1
84.8 85.0 76.9
1.0 1.1 1.2
29.8 28.5 42.9
2.4 2.8 2.4
66.8 67.6 53.5
.... .... ....
12.2 13.3 18.1
4,4 4.0 6.6
83,4 82.7 74.7
P--pa!mitic
IV TABLE A c i d C o m p o s i t i o n of A n i m a l
U
45.5 46.7 51.9
acid;
1.0
59.9
1.7 1.7 2.2
M--myristic
1.1 2.5
5.8
acid
1.0
........ . . . . . . .
29.8
P--palmitic
3.8 3.7
. . . . . . . . . . . . . . . . . . . . . . .
...
TABLE III A c i d C o m p o s i t i o n of V e g e t a b l e M
U--unsaturated S--stearic acid.
acid;
Fats
40
Possible glycerides Human Found ....................................................... Cale ............................................................ Dog F o u n d ...................................................... Calc .......................................................... Ground squirrel Found ....................................................... Calc ............................................................ Chicken Found ........................................................ Calc ............................................................ Pig Found ......................................................... Calc ........................................................... Rabbit Found ......................................................... Calc ............................................................. Guinea pig Found ......................................................... Calc .............................................................
I of X l e g e t a b l e
VOL. 41
SOCIETY
38
Glyceride Carbon
CHEMISTS'
36
Possible glycerides Palm oil Found ........................................................... Calc ............................................................. Linseed oil Found ........................................................... Cale .............................................................. Corn oil Found Calc ........................................................... Cocoa butter Found .......................................................... Calc ............................................................. O l i v e oil F o u n d ............................................................ Calc .............................................................. Cottonseed oil Found .......................................................... Calc .............................................................. Soybean oil Found ........................................................... Calc ..............................................................
0IL
acid
Human Original ............................ Calc ................................ 1 - 3 p o s i t i o n s ................... Dog Original ............................. Calc ................................... 1-3 positions .................. Ground squirrel Original ............................. Calc ............................. 1--3 p o s i t i o n s .................. Chicken Original ............................. Calc ............................ 1 - 3 p o s i t i o n s ................... Pig Original ............................. Calc .......................... 1--3 p o s i t i o n s ................... l~abbit Original .............................. Calc .................................. 1 - 3 p o s i t i o n s .................... Guinea pig Original ............................. Calc .................................. 1-3 positions ........ and
fats, p a r t i c u l a r l y for those of c o m m e r c i a l interest. 0 n l y the i n d i v i d u a l s a t u r a t e d acids have been considered here, the u n s a t u r a t e d acids b e i n g considered as a group. T h a t the d i s t r i b u t i o n of i n d i v i d u a l u n s a t u r a t e d acids m a y also be c a l c u l a t e d i n this w a y is i n d i c a t e d b y a p r e v i o u s p u b l i c a t i o n (4) where the d i s t r i b u t i o n of each i n d i v i d u a l acid was determ i n e d for two fats.
U--unsaturated S--stearic acid.
Technical
1. 2. 3. 4.
acids;
assistance
Fats
M
P
S
U
2.4 2.0 2.4
23.4 25.3 28.5
5.2 4.6 6.8
69.0 68.1 62.3
4.2 3.8 2.7
22.8 24.0 22.9
11.7 11.3 16.5
61.0 60.9 58.2
1.0 1.0 0.9
14.8 14.2 20.1
1.5 1.5 2.0
82.7 83.3 77.0
1.1 1.0 1.1
23.6 24.0 28.9
6.2 5.1 9.0
69,1 70.1 61.0
1.5 l.O 0,7
26.0 25.8 8.2
12.5 13.8 18.9
60,0 59.4 72.2
3.2 3.5 1.8
24.6 24.0 27.4
5.3 5.5 7.8
66.9 67.0 63.0
.... ....
24.0 23.6 27.3
5.9 6.9 8.5
70.1 69.5 64.2
M--myristic
acid;
ACKNOWLEDGMENTS throughout the investigation
P--palmitic
by D. L. McPhee.
REFERENCES Y o u n g s , C. G., a n d M . I t . S u b b a r a m , JAOCS 41, 218 VanderWal, R. J., Ibld. 37, 18~(1960). Y o u n g s , C. G., I b i d . 38, 62 (1961). Subbaram, M. It., and C. G. Youngs, Ibid., in press. [Received
March
17,
1964--Accepted
acid
May
(1964).
20, 1964]
and
