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Relation of Color and Ascorbic Acid to Flavor in Milk from Individual Cows1
R E L A T I O N O F COLOR AND A S C O R B I C A C I D TO F L A V O R I N M I L K F R O M I N D I V I D U A L COWS 1 O. F. GARRETT, H. H. TUCKER, AND F. C. BUTTON New Jersey Agricultural Experiment Station, New ~runsw~cl~, N. J.
I n 1935, Chilson (4) reported that vitamin C (ascorbic acid) exerted a pronounced influence in preventing the development of oxidized or tallowy flavor in milk when it was added directly to milk. Sharp, T r o u t and Guthrie (6) later studied this problem and reported that, " T h e r e is a positive correlation between the rate of oxidation of ascorbic acid and the rate of development of the oxidized flavor." Brown, T h u r s t o n and Dustman (3) f o u n d that, " D r y feeding increased the tendency for oxidized flavor to develop in milk, and grazing on fresh pasture decreased this t e n d e n c y . " These latter findings are corroborated by experiments carried out at the New J e r s e y Agricultural E x p e r i m e n t Station (5). Although it is generally believed that vitamin C in milk cannot be greatly increased by feeding, Brown, Thurston and Dustman (3) f o u n d that the feeding of tomato or lemon juice or pure crystalline ascorbic acid greatly decreased the tendency for oxidized flavor to develop even when the cows were on d r y feed. Anderson (1, 2), in collaboration with H a r b e n b e r g h and Wilson, has reported that " a l l available data concerning feeds strongly suggest a relation between carotene and ability to produce milk of good flavor." I n an experiment conducted at the Sussex branch of the New Jersey Agricultural E x p e r i m e n t Station from December 1, 1936, to April 12, 1937, the following factors were studied: 1. The relation of the percentage of f a t to yellow color of milk. 2. The relation of yellow color to ascorbic acid in milk. 3. The relation of yellow color to flavor of milk. 4. The relation of ascorbic acid to flavor of milk. A total of 28 cows consisting of 16 pure bred Guernseys and 12 pure bred Holsteins were used in this s t u d y which was carried over a period of approximately f o u r months. The first samples were taken on J a n u a r y 7, 1937, and the last ones on April 29, 1937. Results on samples taken on F e b r u a r y 4th and F e b r u a r y 11th are not presented in this study since the milks were inadvertently exposed to bright sunlight before reaching the laboratory. Individual samples from single milkings were taken at weekly intervals and were immediately analyzed for fat b y the Babcock method and for 1 Recently it has been indicated that dehydroascorbic acid shows vitamin C potency. Since the dye-titration method used in this study does not determine the dehydro form the term '' ascorbic acid" is used instead of the term ' 'vitamin C.'' Journal Series paper of the New Jersey Agricultural Experiment Station, department of dairy husbandry. Received for publication November 17, 1937. 121
122
O. F. GARRET% H. H. TUCKER AND F. C. BUTTON
yellow color with the " l a c t o c h r o m e t e r . ''~ Each sample was divided into three parts, the last two of which were stored in glass bottles in a refrigerator at about 40 ° F. The first p a r t of each sample was analyzed for ascorbic acid by the dye-titration method and immediately thereafter was scored for flavor. The second and third parts of each sample were removed from storage on the two succeeding days respectively and were also analyzed for ascorbic acid and scored for flavor. TABLE 1
Showingtherankofpercentageoffat, flavor, aseorbivaoid, andcolor Cow No. 470 428 471 267 467 479 179 465 78 474 273 422 165 92 484 215 478 217 229 276 253 234 284 230 286 431 220 285
G G G G G G G G H G G G G H G H G H H H H I-I H H H G 1:[ G
P e r cent fat ~ rank 6 19 8 7 2 3 5 1 23 12 1O 9 4 17 11 13 15 28 18 24 20 27 25 26 21 13 22 15
Flavor rank ~ F i r s t day 1 2.5 2.5 5 5 5 7.5 7.5 9 11.5 11.5 11.5 11.5 14 15 16.5 16.5 18 19.5 19.5 21 22 23 25 25 25 27 28
T h i r d day 8 2 4 5 6.5 9.5 1 9.5 19.5 6.5 11.5 13 14 15 21 3 16 9.5 17.5 17.5 22 19.5 24.5 23 26 27 24.5 28
Ascorbi¢ acid r a n k S F i r s t day
9
14 13 4 21 3 8 1 5 17 2 10 6 16 22 11 7 19 12 20 18 26 27 23 24 15 25 28
T h i r d day
I
I
5
12 15 4 24 10 9 1 3 19 2 11 6 14 20 8 7 18 13 16.5 16.5 25 22 21 26 23 27 28
Color ~ rank 2 9.5 7 3 8 17 5 1 20 14 6 11 4 18 13 15 12 24 92 19 23 28 26 27 21 9.5 25 16
G---Guernsey; H - - H o l s t e i n . The r a n k i n g of each f a c t o r is based on the average f o r each cow t h r o u g h o u t the experiment f o r t h a t p a r t i c u l a r factor.
In studying the relation between color, fat, ascorbic acid and flavor statistical analyses of the results were made. The coefficients of correlation were obtained in all cases by S p e a r m a n ' s rank-difference method. A correlation coefficient of .4 or better was considered significant. 2 The ' ~L a c t o c h r o m e t e r ' ' is a modification of the P f u n d i n s t r u m e n t devised b y the D e p a r t m e n t of D a i r y H u s b a n d r y of the N e w J e r s e y College of A g r i c u l t u r e in cooperation w i t h the A m e r i c a n Guernsey Cattle Club f o r the r a p i d d e t e r m i n a t i o n of yellow color in milk f r o m all sources.
ASCORBIC ACID A N D
TABLE
123
M I L K FLAVOR
2
Correlations Variables 1.
2. 3.
4. 5. 6. 7.
Color a. b. c. Color Color a. b. c. Color First First Third
Correlation
vs. Percentage of fat A l l s a m p l e s ................................................................................. G u e r n s e y s ....................................................................................... H o l s t e i n s ....................................................................................... vs. F i r s t d a y a s c o r b i e a c i d .......................................... vs. l ~ i r s t d a y f l a v o r A l l s a m p l e s . ................................................................................ G u e r n s e y s ....................................................................................... H o l s t e i n s ..................................................................................... vs. T h i r d d a y f l a v o r ............................................................ d a y a s c o r b i c a c i d vs. F i r s t d a y f l a v o r ............ d a y a s c o r b i c a c i d vs. T h i r d d a y f l a v o r ............ d a y a s c o r b i e a c i d vs. T h i r d d a y f l a v o r ............
coefficient
+.8702 +.5317 +.7366 +.6847 +.7339 +.4834 +.7229 +.6039 +.6996 + .5779 +.5843
RESULTS
There was a close relationship between the percentage of f a t and the color of milk, Tables 1 and 2. This is shown by a correlation coefficient of +.8702 between these two variables when the data from all 28 cows were used in the computation. When the data were segregated as to breeds the correlation coefficients were + .5317 and + .7366 respectively for Guernseys and Holsteins. This indicates that the percentage of f a t had a more positive effect on the color of the Holstein milk than it had on the Guernsey milk. The correlation coefficient of + .6847 between color and first day ascorbic acid (ascorbic acid determined on the day of the milking) indicates a very significant relationship between these two variables. I t is quite possible that both factors are not only closely related to feeds and feeding practices but also may depend alike on certain h e r e d i t a r y factors of the individual cow and on the physiological condition of the cow. The yellow color of milk is primarily due to the yellow pigment, carotene. A close inspection of the molecular structure and the reactions of carotene reveals that it has reducing properties. I t is known that the presence of reducing substances in milk will tend to prevent an oxidizing reaction in the milk. I t would be expected, therefore, that carotene would exert a protective action on the flavor of milk. That a close positive relationship exists is shown by correlation coefficients of + .7339 between color and first day flavor and + 6039 between color (first day) and third day flavor (scored after about 56 hours' storage). These two coefficients indicate that, while the relationships between the two factors" may be p a r t l y coincidental, color or carotene does help to stabilize flavor. This is especially indicated by the correlation between color and flavor a f t e r the milk has been in storage for some considerable time. Ever since the molecular structure of ascorbic acid was discovered, it has been known that the compound has reducing properties. Accordingly
~t b~
TABLE 3
Distribution of flavor scares in aseorbiv avid c~asses Ascorbic a c i d class intervals, rag./1. 30 27 24 21 18 15 12
............... ............... ............... ............... ............. ............... ...............
0
...............
6
...............
3
...............
0
...............
T o t a l s .........
Number
F l a v o r score class i n t e r v a l s
eases
23
22.5
3 4 29 62 138 121 92 34
1
496
8 24 30 48 58 19 12 1 202
22
21.5
I
21
20.5
20
i n each class
for each class 22.88 22.80 22.85 22.72 22.68 22.42 22.20 21.73 21.45 21.34
6
14 25 7 5 1
4 8 11 11 5 3
2 6 12 5 2 1
10 5 2
114
69
42
28
26
981
4
13
score
19.5-12
4 5 39 100 202 237 230 lOO 48 16
1 2 10 14 36 26
Average
P e r cen t o f cases w i t h score of 23
P e r cen t o f cases w i t h score below 21.5
75.0 80.0 74.4 62.0 68.3 40.0 34.0 22.9 12.5
0 0 0 0 3.5 7.2 12.6 28.0 25.0 37.5
50.6
10.2
51.1
Q
ASCORBIC ACID AND MILK FLAVOR
125
it can be predicted that ascorbic acid should exert a protective action on the flavor of milk similar to that shown by carotene. A statistical analysis of the data from all 28 cows shows that such a relationship does e x i s t . The correlation coefficient of + .6996 between first day ascorbic acid and first day flavor is quite significant, while significant coefficients between first day ascorbic acid and third day flavor and between third day ascorbie acid and third day flavor indicate that ascorbic acid helps to stabilize flavor. This relationship is further shown in Table 3. In making this table the ascorbic acid values were divided into three-milligram intervals; the flavor scores were divided into 0.5-point intervals. In plotting the distribution, only data from single samples were used; no averages appear in this table. There was a total of 981 samples on which the flavor score and ascorbic acid content were obtained. An inspection of the average flavor score for each ascorbic acid class interval reveals that with a decrease in the amount of aseorbic acid a decrease in the flavor score occurs. A furt her inspection of the data reveals that an apparent critical point in the relation of ascorbic acid to good flavor lies somewhere between 15 and 18 milligrams of aseorbic acid per liter of milk. CONCLUSIONS
From the data presented in this paper it can be concluded that in the milk from individual cows: 1. There is a close relation between percentage of fat and yellow color. 2. There is a significant association of yellow color and ascorbic acid. 3. High carotene and high ascorbic acid are coincidental to and help to preserve good flavor in milk. I t is recommended, therefore, that special efforts be made to preserve the carotene in roughages and that winter feeding practices be followed so as to maintain a high level of yellow color in the milk. I t is further recommended that special efforts be made to preserve the ascorbic acid in milk by subjecting the milk to a minimum amount of aeration, by prevention of contamination with such metals as copper, iron and nickel, and by protection of the milk from sunlight. ACKNOVq]0EDGMENT
The authors wish to thank l~r. Carl de Boer and Mr. John Mixner, graduate students, for much of the laboratory work involved in this study. REFERENCES
(1) ANDERSON~ J. A., "[-TARDRNBERG,J. G.~ AND WlLSON~ L . T . Concerning the cause of rancid and oxidized flavors of bovine origin. JouR. DAIRY ScI. 19. 483. 1936. (2) ANDERSON, J . A . Off-flavored milk. Milk Dealer 26: 60. 1936. (3) BROWN, W. C., THURSTON, L. M., AND DUSTMAN, 1~. B. Oxidized flavor in milk. IV. Studies of the relation of feed of the cow to oxidized flavor. JOUR. DAIRY ScI. 20: 133. 1937.
126
O. F. GARRET% H. It. TUCKER AND F. C. BUTTON
(4) CHIT.SON,W . H . What causes most common off:flavors of market milk~ Milk Plant l~onthly 24 : 24 ; 30. 1935. (5) GARRETT, O . F . Cardboard flavor. New Jersey Agriculture 19: 3. 1937. (6) SHARP, P. F.~ TROUT, G. ~J~.~ AND GUTHRIE, E. S. Vitamin C~ copper and the oxidized flavor of milk. Mfll~ P l a n t Monthly 26: 32. 1937.
I n 1935, Chilson (4) reported that vitamin C (ascorbic acid) exerted a pronounced influence in preventing the development of oxidized or tallowy flavor in milk when it was added directly to milk. Sharp, T r o u t and Guthrie (6) later studied this problem and reported that, " T h e r e is a positive correlation between the rate of oxidation of ascorbic acid and the rate of development of the oxidized flavor." Brown, T h u r s t o n and Dustman (3) f o u n d that, " D r y feeding increased the tendency for oxidized flavor to develop in milk, and grazing on fresh pasture decreased this t e n d e n c y . " These latter findings are corroborated by experiments carried out at the New J e r s e y Agricultural E x p e r i m e n t Station (5). Although it is generally believed that vitamin C in milk cannot be greatly increased by feeding, Brown, Thurston and Dustman (3) f o u n d that the feeding of tomato or lemon juice or pure crystalline ascorbic acid greatly decreased the tendency for oxidized flavor to develop even when the cows were on d r y feed. Anderson (1, 2), in collaboration with H a r b e n b e r g h and Wilson, has reported that " a l l available data concerning feeds strongly suggest a relation between carotene and ability to produce milk of good flavor." I n an experiment conducted at the Sussex branch of the New Jersey Agricultural E x p e r i m e n t Station from December 1, 1936, to April 12, 1937, the following factors were studied: 1. The relation of the percentage of f a t to yellow color of milk. 2. The relation of yellow color to ascorbic acid in milk. 3. The relation of yellow color to flavor of milk. 4. The relation of ascorbic acid to flavor of milk. A total of 28 cows consisting of 16 pure bred Guernseys and 12 pure bred Holsteins were used in this s t u d y which was carried over a period of approximately f o u r months. The first samples were taken on J a n u a r y 7, 1937, and the last ones on April 29, 1937. Results on samples taken on F e b r u a r y 4th and F e b r u a r y 11th are not presented in this study since the milks were inadvertently exposed to bright sunlight before reaching the laboratory. Individual samples from single milkings were taken at weekly intervals and were immediately analyzed for fat b y the Babcock method and for 1 Recently it has been indicated that dehydroascorbic acid shows vitamin C potency. Since the dye-titration method used in this study does not determine the dehydro form the term '' ascorbic acid" is used instead of the term ' 'vitamin C.'' Journal Series paper of the New Jersey Agricultural Experiment Station, department of dairy husbandry. Received for publication November 17, 1937. 121
122
O. F. GARRET% H. H. TUCKER AND F. C. BUTTON
yellow color with the " l a c t o c h r o m e t e r . ''~ Each sample was divided into three parts, the last two of which were stored in glass bottles in a refrigerator at about 40 ° F. The first p a r t of each sample was analyzed for ascorbic acid by the dye-titration method and immediately thereafter was scored for flavor. The second and third parts of each sample were removed from storage on the two succeeding days respectively and were also analyzed for ascorbic acid and scored for flavor. TABLE 1
Showingtherankofpercentageoffat, flavor, aseorbivaoid, andcolor Cow No. 470 428 471 267 467 479 179 465 78 474 273 422 165 92 484 215 478 217 229 276 253 234 284 230 286 431 220 285
G G G G G G G G H G G G G H G H G H H H H I-I H H H G 1:[ G
P e r cent fat ~ rank 6 19 8 7 2 3 5 1 23 12 1O 9 4 17 11 13 15 28 18 24 20 27 25 26 21 13 22 15
Flavor rank ~ F i r s t day 1 2.5 2.5 5 5 5 7.5 7.5 9 11.5 11.5 11.5 11.5 14 15 16.5 16.5 18 19.5 19.5 21 22 23 25 25 25 27 28
T h i r d day 8 2 4 5 6.5 9.5 1 9.5 19.5 6.5 11.5 13 14 15 21 3 16 9.5 17.5 17.5 22 19.5 24.5 23 26 27 24.5 28
Ascorbi¢ acid r a n k S F i r s t day
9
14 13 4 21 3 8 1 5 17 2 10 6 16 22 11 7 19 12 20 18 26 27 23 24 15 25 28
T h i r d day
I
I
5
12 15 4 24 10 9 1 3 19 2 11 6 14 20 8 7 18 13 16.5 16.5 25 22 21 26 23 27 28
Color ~ rank 2 9.5 7 3 8 17 5 1 20 14 6 11 4 18 13 15 12 24 92 19 23 28 26 27 21 9.5 25 16
G---Guernsey; H - - H o l s t e i n . The r a n k i n g of each f a c t o r is based on the average f o r each cow t h r o u g h o u t the experiment f o r t h a t p a r t i c u l a r factor.
In studying the relation between color, fat, ascorbic acid and flavor statistical analyses of the results were made. The coefficients of correlation were obtained in all cases by S p e a r m a n ' s rank-difference method. A correlation coefficient of .4 or better was considered significant. 2 The ' ~L a c t o c h r o m e t e r ' ' is a modification of the P f u n d i n s t r u m e n t devised b y the D e p a r t m e n t of D a i r y H u s b a n d r y of the N e w J e r s e y College of A g r i c u l t u r e in cooperation w i t h the A m e r i c a n Guernsey Cattle Club f o r the r a p i d d e t e r m i n a t i o n of yellow color in milk f r o m all sources.
ASCORBIC ACID A N D
TABLE
123
M I L K FLAVOR
2
Correlations Variables 1.
2. 3.
4. 5. 6. 7.
Color a. b. c. Color Color a. b. c. Color First First Third
Correlation
vs. Percentage of fat A l l s a m p l e s ................................................................................. G u e r n s e y s ....................................................................................... H o l s t e i n s ....................................................................................... vs. F i r s t d a y a s c o r b i e a c i d .......................................... vs. l ~ i r s t d a y f l a v o r A l l s a m p l e s . ................................................................................ G u e r n s e y s ....................................................................................... H o l s t e i n s ..................................................................................... vs. T h i r d d a y f l a v o r ............................................................ d a y a s c o r b i c a c i d vs. F i r s t d a y f l a v o r ............ d a y a s c o r b i c a c i d vs. T h i r d d a y f l a v o r ............ d a y a s c o r b i e a c i d vs. T h i r d d a y f l a v o r ............
coefficient
+.8702 +.5317 +.7366 +.6847 +.7339 +.4834 +.7229 +.6039 +.6996 + .5779 +.5843
RESULTS
There was a close relationship between the percentage of f a t and the color of milk, Tables 1 and 2. This is shown by a correlation coefficient of +.8702 between these two variables when the data from all 28 cows were used in the computation. When the data were segregated as to breeds the correlation coefficients were + .5317 and + .7366 respectively for Guernseys and Holsteins. This indicates that the percentage of f a t had a more positive effect on the color of the Holstein milk than it had on the Guernsey milk. The correlation coefficient of + .6847 between color and first day ascorbic acid (ascorbic acid determined on the day of the milking) indicates a very significant relationship between these two variables. I t is quite possible that both factors are not only closely related to feeds and feeding practices but also may depend alike on certain h e r e d i t a r y factors of the individual cow and on the physiological condition of the cow. The yellow color of milk is primarily due to the yellow pigment, carotene. A close inspection of the molecular structure and the reactions of carotene reveals that it has reducing properties. I t is known that the presence of reducing substances in milk will tend to prevent an oxidizing reaction in the milk. I t would be expected, therefore, that carotene would exert a protective action on the flavor of milk. That a close positive relationship exists is shown by correlation coefficients of + .7339 between color and first day flavor and + 6039 between color (first day) and third day flavor (scored after about 56 hours' storage). These two coefficients indicate that, while the relationships between the two factors" may be p a r t l y coincidental, color or carotene does help to stabilize flavor. This is especially indicated by the correlation between color and flavor a f t e r the milk has been in storage for some considerable time. Ever since the molecular structure of ascorbic acid was discovered, it has been known that the compound has reducing properties. Accordingly
~t b~
TABLE 3
Distribution of flavor scares in aseorbiv avid c~asses Ascorbic a c i d class intervals, rag./1. 30 27 24 21 18 15 12
............... ............... ............... ............... ............. ............... ...............
0
...............
6
...............
3
...............
0
...............
T o t a l s .........
Number
F l a v o r score class i n t e r v a l s
eases
23
22.5
3 4 29 62 138 121 92 34
1
496
8 24 30 48 58 19 12 1 202
22
21.5
I
21
20.5
20
i n each class
for each class 22.88 22.80 22.85 22.72 22.68 22.42 22.20 21.73 21.45 21.34
6
14 25 7 5 1
4 8 11 11 5 3
2 6 12 5 2 1
10 5 2
114
69
42
28
26
981
4
13
score
19.5-12
4 5 39 100 202 237 230 lOO 48 16
1 2 10 14 36 26
Average
P e r cen t o f cases w i t h score of 23
P e r cen t o f cases w i t h score below 21.5
75.0 80.0 74.4 62.0 68.3 40.0 34.0 22.9 12.5
0 0 0 0 3.5 7.2 12.6 28.0 25.0 37.5
50.6
10.2
51.1
Q
ASCORBIC ACID AND MILK FLAVOR
125
it can be predicted that ascorbic acid should exert a protective action on the flavor of milk similar to that shown by carotene. A statistical analysis of the data from all 28 cows shows that such a relationship does e x i s t . The correlation coefficient of + .6996 between first day ascorbic acid and first day flavor is quite significant, while significant coefficients between first day ascorbic acid and third day flavor and between third day ascorbie acid and third day flavor indicate that ascorbic acid helps to stabilize flavor. This relationship is further shown in Table 3. In making this table the ascorbic acid values were divided into three-milligram intervals; the flavor scores were divided into 0.5-point intervals. In plotting the distribution, only data from single samples were used; no averages appear in this table. There was a total of 981 samples on which the flavor score and ascorbic acid content were obtained. An inspection of the average flavor score for each ascorbic acid class interval reveals that with a decrease in the amount of aseorbic acid a decrease in the flavor score occurs. A furt her inspection of the data reveals that an apparent critical point in the relation of ascorbic acid to good flavor lies somewhere between 15 and 18 milligrams of aseorbic acid per liter of milk. CONCLUSIONS
From the data presented in this paper it can be concluded that in the milk from individual cows: 1. There is a close relation between percentage of fat and yellow color. 2. There is a significant association of yellow color and ascorbic acid. 3. High carotene and high ascorbic acid are coincidental to and help to preserve good flavor in milk. I t is recommended, therefore, that special efforts be made to preserve the carotene in roughages and that winter feeding practices be followed so as to maintain a high level of yellow color in the milk. I t is further recommended that special efforts be made to preserve the ascorbic acid in milk by subjecting the milk to a minimum amount of aeration, by prevention of contamination with such metals as copper, iron and nickel, and by protection of the milk from sunlight. ACKNOVq]0EDGMENT
The authors wish to thank l~r. Carl de Boer and Mr. John Mixner, graduate students, for much of the laboratory work involved in this study. REFERENCES
(1) ANDERSON~ J. A., "[-TARDRNBERG,J. G.~ AND WlLSON~ L . T . Concerning the cause of rancid and oxidized flavors of bovine origin. JouR. DAIRY ScI. 19. 483. 1936. (2) ANDERSON, J . A . Off-flavored milk. Milk Dealer 26: 60. 1936. (3) BROWN, W. C., THURSTON, L. M., AND DUSTMAN, 1~. B. Oxidized flavor in milk. IV. Studies of the relation of feed of the cow to oxidized flavor. JOUR. DAIRY ScI. 20: 133. 1937.
126
O. F. GARRET% H. It. TUCKER AND F. C. BUTTON
(4) CHIT.SON,W . H . What causes most common off:flavors of market milk~ Milk Plant l~onthly 24 : 24 ; 30. 1935. (5) GARRETT, O . F . Cardboard flavor. New Jersey Agriculture 19: 3. 1937. (6) SHARP, P. F.~ TROUT, G. ~J~.~ AND GUTHRIE, E. S. Vitamin C~ copper and the oxidized flavor of milk. Mfll~ P l a n t Monthly 26: 32. 1937.