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The availability of lysine from Torula yeast
ARCHIVES
OF BIOCHEMISTRY
AND
The Availability
BIOPHYSICS
71,
414-422 (1957)
of Lysine from Torula Yeast1
Wei S. Tsien, Elton L. Johnson and Irvin E. Liener From the Departments of Poultry Husbandry and AgricuWural University of Minnesota, St. Paul, Minnesota Received February
Biochemistry,
18, 1957
INTRODUCTION
Among the more important factors governing the choice of a supplementary protein for lysine-deficient cereal proteins used in human, livestock, and poultry feeding are primarily its lysine content, and, from a practical viewpoint, its cost and availability. On both of these counts Torula yeast appears to hold considerable promise. Its lysine content compares favorably with that of animal protein (l), and it is potentially capable of being inexpensively produced on a large scale from the waste products of the paper industry (2, 3). It is well recognized that the amino acid analysis of foodstuffs may not, be a reliable index of the nutritive quality of its protein. The biological availability of lysine in particular may diverge appreciably from its analytical value in a number of proteins (4-6). The present studies were undertaken to establish the availability of lysine from To&a yeast with chicks and rats receiving basal diets containing adequate amounts of all the known dietary essentials save lysine. During the course of these studies it became evident that growth response was being complicated by whit appeared to be unidentified nutrients in Torula yeast. More definitive information was sought using nitrogen retention and lysine absorption as criteria of lysine availability. EXPERIMENTAL
The Torula yeast used in this study was a dried flaked products which had been prepared in accordance with published details (7). This sample of yeast had a 1 Paper No. 3689, Scientific Journal Series, Minnesota Agricultural Experiment Station. This work has been supported by grants from the Lake States Yeast Corporation and the Sulphite Pulp Manufacturers’ Research League. 2 Provided by Dr. A. J. Wiley, Sulphite Pulp Manufacturers’ Research League, Appleton, Wisconsin. 414
AVAILABILITY
OF
YEAST
TABLE
Basal Lysine-Deficient
415
LYSINE
I
Ration Used in Chick Studies Per cent of ration
Ingredient Corn-gluten meal (42% protein) Zein Alfalfa meal, dehydrated (17% tein) Soybean oil meal (44y0 protein) Dried fish solubles (12.8% N) Steamed bone meal Corn starch Cerelose Manganese sulfate Iodized sodium chloride Ground limestone L-Arginine.HCl oL-Tryptophan or.-Methionine Glycine Choline chloride (25y0 dry mix)a Vitamin A and D concentrateb Vitamin E Water-soluble vitamin mixd
25.00 6.00 2.00
pro-
1.60 2.50 4.50 27.05 27.05 0.05 0.50 0.60 0.75 0.15 0.25 0.70 0.40 0.60 0.05 0.25
a Merck and Co., Rahway, N. J. * Contains 2250 I. U. vitamin A and 300 I. C. units D,/g. Nopco Chemical Co., Harrison, N. J. c Myvamix. 20,000 I.U./lb., Distillation Products Industries, Rochester, N. Y. d No. 95 Vitamin Mix (Merck) which, according to the manufacturer, contains per lb.: 2 g. riboflavine, 4 g. calcium pantothenate, 20 g. niacin, and 106 g. choline chloride. In addition, the following vitamins were added to give per kg. ration: 2 mg. thiamine.HCl, 3 mg. pyridoxine, 700 fig. folic acid, 90 pg. biotin, and 11 Mg. vitamin BIT protein content of 47.470 (N X 6.25) and a lysine content of 4.1y0. All lysine values reported were determined by microbiological assay using Leuconostoc mesenteroidea P-60 (8). Day-old New Hampshire chicks were divided into nine lots of six each with due regard to weight and sex. One group received the semipractical basal ration shown in Table I. This ration was found to contain 0.23% lysine by actual analysis. Four lots were fed the basal diet supplemented with 0.28,0.48,0.68, and 0.88% L-lysine.HC1.3 Torula yeast was fed to the remaining four groups at levels of 5, 10,15, and 20y0 of the basal diet. The addition of lysine or protein was accompanied by an equivalent deduction in the protein of the basal diet as derived from corngluten meal and zein. The corn starch of the basal ration was reduced to corre3 Ninety-five per mington, Delaware.
cent
pure
product
obtained
from
E. I. DuPont
and
Co.,
Wil-
416
TSIEN,
Composition Mixture A n-Arginine n-Histidine
JOHNSON
AND
TABLE II of Amino Acid Mixtures Amino acid . HCI . HCI . Hz0
L-Leucine nn-Methionine nn-Phenylalanine nn-Threonine nn-Tryptophan nn-Valine nn-Alanine Dr.-Aspartic acid L-Cystine n-Glutamic acid L-Tyrosine Glycine A
13,521
Mixture B nn-Methionine nn-Tryptophan n-Histidine nL-Phenylalanine Total Mixture B Total Mixture A and Mixture
Used in Rat Studies nag./100g. ration 863 451 1729 1644 468 693 1040 241 1872 577 648 334 2450 660 51
DL-ISOhCiDf?
Total Mixture
LIBlNER
300 40 80 140 B
560 14,081
spond to the nonprotein material introduced by the various levels of To&a yeast. Diets and water were provided ad l&turn for 3 weeks during which time weekly weight changes and total food consumption were recorded. At the completion of the experiment, the animals were sacrificed and the carcasses stored in the freezer for nitrogen analysis. Kjeldahl nitrogen was determined on a representative sample of the ground carcass. Nitrogen retention was aalculated as the difference between the N content of each carcass at completion of the experiment, and the N content of the carcasses from day-old chicks sacrificed and frozen at the beginning of the experiment (average of 20 birds, 10 males and 10 females). The difference in N carcass values per unit of body weight before and after experimentation was likewise calculated. Young rats weighing an average of 60 g. were divided into seven groups with six animals equally allotted to each group on the basis of age, sex, and initial weight. One group was fed a basal purified ration containing the amino acid mixtures A and B (14%) shown in Table II, 5% corn oil,’ 4% salt mixture (9), 74% ’ Mazola.
AVAILABILITY
OF YEAST LYSINE
417
corn starch, 1% corn oil-fat soluble vitamin mix, and 2% sucrose-water soluble vitamin mix.6 The latter two components were so formulated as to provide all the known vitamins (9). Three groups of rats were fed the basal ration to which had been added 0.3,0.6, and 0.9% n-lysine .* The remaining three groups were fed the basal ration supplemented with 7,14, and 21% Torula yeast. In order to maintain the nitrogen of the ration at a constant level, the addition of synthetic lysine or protein in the form of Torula yeast was compensated for by an equivalent reduction in amino acid mixture A. The proportion of amino acid mixture B in the basal ration was kept constant to insure an adequate supply of methionine, tryptophan, histidine, and phenylalanine which tends to be marginal in Torula yeast protein (7). Appropriate reductions in corn starch were made corresponding to the nonprotein contribution of the yeast supplement. All rats were individually housed and given food and water ad lib&m. Weight changes and food consumption were recorded for a period of 3 weeks. In the second week of the preceding experiment, two animals from each group were randomly selected for Iysine absorption studies. Feces were quantitatively collected for a period of 11 days. The fecal pellets could be readily separated by hand from any spilled feed which frequently fell beneath the cages. The absorption of lysine was calculated from data based on the lysine content of the ration and the feces in conjunction with the quantity of food consumed and feces excreted during the period of observation. RESULTS
In Fig. 1 is plotted the growth responseof chicks fed the basal ration (Table I) supplemented with graded levels of synthetic lysine or its equivalent in the form of Torulu yeast as indicated by microbiological analysis. Additional data pertaining to feed efficiency and nitrogen retention are presented in Table III. Up to approximately 0.5 % synthetic or natural lysine there appears to be no significant difference6 in the growth responseelicited by synthetic lysine or yeast providing an equivalent level of lysine. Synthetic lysine in excess of 0.68 % produced no further increment in weight gain; the total level of lysine in such a diet is 0.91% which may be compared with the level of 0.9 % recommended by the National Research Council (10). The superiority of Torula yeast at levels of 15 and 20 % over diets providing comparable levels of synthetic lysine is readily apparent from Fig. 1, and its significance was verified by statistical analysis.E Feed efficiency and nitrogen retention on a per-chick basis closely paralleled the pattern of growth response,with the highest levels of yeast ‘Modified to include vitamin Blz at a level of 4 ag./106 g. ration. (LSignificant differences as used hereafter in this paper will imply a “P” value <0.05 calculated from the analysis of variance. Intertreatment comparisons were made from tables published by May (18).
418
TSIEN,
JOHNSON
t I
AND
I I I
LIENER
I , ,
, ‘
0
0.2 0.4 0.6 0.8 1.0 % L- LYSINE ADDED I I I # I , , , , , J 0 5 10 15 20 25 %TORULA YEAST ADDED FIG. 1. Growth responseof chicks to gradedlevels of L-lysine (0 -0)
or
Torda yeast (*---0) added to basal lysine-deficient diet. The lower abcissa referring to levels of Torula yeast has been adjusted to coincide with the upper
abcissabasedon a lysine content of 4% asdeterminedby microbiologicalanalysis. TABLE III Feed Supplement
E&iency to basal
and Nitrogen ration
None
0.28% L-lysine 0.48yo L-lysine 0.68% L-lysine
Data
in Chick Nitrogen g./ckick
6.3 3.7 2.7 2.2 2.4 4.2 2.6 2.2 2.1
0.60 1.87 2.82 3.85 3.70 1.22 3.02 4.11
Studies retentiona #./kg. body
WA
2.7
19.4
4.91
25.4 40.5 55.5 8.3 21.2 35.1 42.1
between initia1 and final values as described
in the text.
0.88% L-lysine 5% To&a yeast 10% Torula yeast 15% Torula yeast 20% To&a yeast = Difference
Retention Feed efficiency g. feed/g. gain
producing a more efficient utilization of the feed and a greater retention of nitrogen than could be accounted for on the basis of its lysine content. When nitrogen retention is expressed on a per unil body weight basis, however, the retention of nitrogen from To&a yeast was consistently
AVAILABILITY
OF
YEAST
LYSINE
419
less than that from the lysine-supplemented diets, the former averaging about 75 % of the latter. If the lysine of Torula yeast is not completely available as the nitrogen retention data would indicate, then animals receiving 15 % yeast should respond to further additions of synthetic lysine. In a separate experiment it was found that chicks receiving the basal diet plus 15 % To&a yeast gained 109 g. in 3 weeks, whereas a similar lot of chicks receiving an additional supplement of 0.2% n-lysine gained 155 g. This would indicate that superior growth of chicks on high levels of Torula yeast can be attained even though the available lysine provided therein may be suboptimal for growth. Rat Studies The results of the growth study with rats are presented in Fig. 2. At a lysine level of 0.6 % or less, the growth curves of rats receiving synthetic lysine or its equivalent in the form of Toruh yeast were superimposable. The 21% level of Torula yeast, however, produced a significantly greater growth response than one would predict on the basis of its lysine content. In common with the experience of others (11,12), protein efficiency
0 I 0
0.2 0.4 0.6 08 1.0 % L-LYSINE ADDED R I1 II I I I fi I1 5 IO 15 20 25 %TORULA YEAST ADDED
FIG. 2. Growth response of rats to graded levels of L-lysine (O0) or To&a yeast (a- - - -@) added to basal lysine-deficient diet. Adjustments of abcissas were made as described in Fig. 1.
420
TSIEN,
Protein Supplement
Eflciency
to basal
ration
JOHNSON
AND
LIENER
TABLE IV and Lysine Absorption Protein
Data in Rat Studies
e5ciency’
Lysine mg./rot/day
None 0.3% L-lysine 0.6% L-lysine 0.9% L-lysine 7% Torula yeast 14yoTot-da yeast 21o/oTort&la yeast
-0.90 0.15 1.34 1.76 0.13 1.17 2.03
0 43 116 169 33 110 169
0 Grams gain in wt./g. protein consumed. b Milligrams lysine ingested - (mg. lysine excreted basal diet). c (Lysine absorbed/lysine ingested) X 100.
absorbedb
n&/g. body wt./day
%c
0 0.67 1.21 1.67 0.54 1.17 1.39
99.5 98.9 99.9 91.4 90.9 90.2
- mg. lysine excreted on
values (Table IV) were found to parallel growth response, and also showed a significant difference at the highest level of lysine and Torula yeast supplementation in favor of the latter. With reference to the lysine absorption data in Table IV, the same total quantities of lysine were absorbed from both the synthetic and yeast-supplemented diets at essentially equivalent levels of lysine. In proportion to body weight, however, the absorption of lysine from the Torula yeast diets were consistently inferior to the synthetic diets and averaged about 88 % of the latter. In relation to the amount of lysine ingested, the yeast lysine was about 90% as efficiently absorbed as the synthetic amino acid. The fact that the rats receiving the yeast diets absorbed the same total quantity of lysine as those fed the synthetic diets, in spite of the decreased availability of this amino acid from the former, is most likely due to the consumption of greater amounts of food, a logical consequence of enhanced growth. DISCUSSION
Although the primary object of the present study was to evaluate the availability of lysine from Torula yeast, the growth responseswhich were obtained were not solely attributable to this factor. The measurement of lysine availability on the basis of growth alone (or values calculated therefrom such as feed or protein efficiencies), as proposed by Schweigert and Guthneck (5), could not therefore be applied to Toruh yeast. Thus, from the growth curves of Figs. 1 and 2, one would be forced
AVAILABILITY
OF
YEAST
LYSINE
421
to conclude that at lower levels of Torula yeast the lysine was completely available; whereas at the higher levels of yeast intake the impossible conclusion would be reached that the availability of lysine exceeded 100 %. On the other hand, measurements involving nitrogen retention in chicks and lysine absorption in rats, expressed on the basis of unit body weight, revealed that 75 and 90 % of the lysine of Torulu yeast was available to the chick and rat, respectively. In spite of the fact that the availability of yeast lysine was somewhat less than that of the synthetic product, greater growth responses were obtained with both chicks and rats at levels of Torula yeast ranging from 15 to 20 % of the diet. Since this enhanced growth was observed at levels of yeast calculated to provide lysine sufficient for optimal growth on the purified diets, unrecognized growth factors may well be implicated here. It is also possible that the substitution of yeast protein for the protein of corn-gluten meal and zein (in the chick experiments) or amino acid mixture h (in the rat experiments) may have produced a more favorable balance with respect to amino acids or minerals. The curious finding that the retention of nitrogen per unit body weight of the more rapidly growing chicks fed Torula yeast was less than that of their counterparts receiving synthetic lysine may be due to the preferential deposition of a nonprotein constituent such as fat, an effect similar to that produced by vitamin Blz on the body composition of the rat (13). In addition to Torula yeast being a rich source of the vitamin B complex (14)) these studies further establish the unique value of Torula yeast as a protein supplement and a possible source of unidentified growth factors. However, in view of the adverse physiological effects which high levels of yeast are known to produce, such as liver damage (15)) calciumphosphorus imbalance (16)) and methionine deficiency (17)) further study is warranted before the extensive use of Torula yeast can be recommended.
SUMMARY The availability of lysine from Tom.@ yeast has been measured with chicks and rats on the basis of growth response, feed or protein efficiency, nitrogen retention (chicks), and lysine absorption (rats). Consistently greater growth responses were obtained on lysine-deficient diets supplemented with 15-20 % Torula yeast than could be accounted for on the basis of the lysine so provided. Measurements involving weight gains,
422
TSIEN,
JOHNSON
AND
LIENER
therefore, did not prove to be adequate criteria for evaluating lysine availability. On the other hand, nitrogen retention data disclosed that yeast lysine was about 75 % as available to the chick as the synthetic substance. With rats, yeast lysine was about 90 % as efficiently absorbed as the synthetic amino acid. REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17, 18.
BLOCK, R. J., AND MITCHELL, H. H., Nutrition Abstr. and Rev. 16,249 (1946-47). Anonymous, Nutrition Rev. 7, 86 (1949). WHITE, J., “Yeast Technology,” p. 302. Chapman and Hall, London, 1954. KUIKEN, K. A., AND LYMAN, L. M., J. Nutrition 36,359 (1948). SCHWEIGERT, B. S., AND GUTHNECK, B. T., J. Nutrition 49, 277 (1953). GUTHNECK, B. T., AND SCHWEIGERT, B. S., J. Nutrition 49, 289 (1953). INSKEEP, G. C., WILEY, A. J., HOLDERBY, J. M., AND HUGHES, L. P., Ind. Eng. Chem. 43, 1702 (1951). SNELL, E. E., Advances in Protein Chem. 2, 85 (1945). SCHULTZE, M. O., J. Nutrition 41, 103 (1950). National Research Council, “Nutrient Requirements for Poultry.!! Natl. Research Council Publ. 301, 27 (1954). HEGSTED, D. M., AND WORCESTER, J., J. Nutrition 33,685 (1947). SHERWOOD, F. W., AND WELDON, V., J. Nutrition 49, 153 (1953). LING, C. T., AND CHOW, B. F., J. Biol. Chem. 198, 439 (1952). DUNN, C. G., Wallerstein Labs. Communs. 16, 61 (1952). SCHWARZ, K., Ann. N. Y. Acad. 2%. 67, 615 (1954). THAYSEN, A. C., Food 14, 116 (1945). KLOSE, A. A., AND FEVOLD, H. L., J. Nutrition 29, 421 (1945). MAY, J. M., Biometrika 39, 192 (1952).
OF BIOCHEMISTRY
AND
The Availability
BIOPHYSICS
71,
414-422 (1957)
of Lysine from Torula Yeast1
Wei S. Tsien, Elton L. Johnson and Irvin E. Liener From the Departments of Poultry Husbandry and AgricuWural University of Minnesota, St. Paul, Minnesota Received February
Biochemistry,
18, 1957
INTRODUCTION
Among the more important factors governing the choice of a supplementary protein for lysine-deficient cereal proteins used in human, livestock, and poultry feeding are primarily its lysine content, and, from a practical viewpoint, its cost and availability. On both of these counts Torula yeast appears to hold considerable promise. Its lysine content compares favorably with that of animal protein (l), and it is potentially capable of being inexpensively produced on a large scale from the waste products of the paper industry (2, 3). It is well recognized that the amino acid analysis of foodstuffs may not, be a reliable index of the nutritive quality of its protein. The biological availability of lysine in particular may diverge appreciably from its analytical value in a number of proteins (4-6). The present studies were undertaken to establish the availability of lysine from To&a yeast with chicks and rats receiving basal diets containing adequate amounts of all the known dietary essentials save lysine. During the course of these studies it became evident that growth response was being complicated by whit appeared to be unidentified nutrients in Torula yeast. More definitive information was sought using nitrogen retention and lysine absorption as criteria of lysine availability. EXPERIMENTAL
The Torula yeast used in this study was a dried flaked products which had been prepared in accordance with published details (7). This sample of yeast had a 1 Paper No. 3689, Scientific Journal Series, Minnesota Agricultural Experiment Station. This work has been supported by grants from the Lake States Yeast Corporation and the Sulphite Pulp Manufacturers’ Research League. 2 Provided by Dr. A. J. Wiley, Sulphite Pulp Manufacturers’ Research League, Appleton, Wisconsin. 414
AVAILABILITY
OF
YEAST
TABLE
Basal Lysine-Deficient
415
LYSINE
I
Ration Used in Chick Studies Per cent of ration
Ingredient Corn-gluten meal (42% protein) Zein Alfalfa meal, dehydrated (17% tein) Soybean oil meal (44y0 protein) Dried fish solubles (12.8% N) Steamed bone meal Corn starch Cerelose Manganese sulfate Iodized sodium chloride Ground limestone L-Arginine.HCl oL-Tryptophan or.-Methionine Glycine Choline chloride (25y0 dry mix)a Vitamin A and D concentrateb Vitamin E Water-soluble vitamin mixd
25.00 6.00 2.00
pro-
1.60 2.50 4.50 27.05 27.05 0.05 0.50 0.60 0.75 0.15 0.25 0.70 0.40 0.60 0.05 0.25
a Merck and Co., Rahway, N. J. * Contains 2250 I. U. vitamin A and 300 I. C. units D,/g. Nopco Chemical Co., Harrison, N. J. c Myvamix. 20,000 I.U./lb., Distillation Products Industries, Rochester, N. Y. d No. 95 Vitamin Mix (Merck) which, according to the manufacturer, contains per lb.: 2 g. riboflavine, 4 g. calcium pantothenate, 20 g. niacin, and 106 g. choline chloride. In addition, the following vitamins were added to give per kg. ration: 2 mg. thiamine.HCl, 3 mg. pyridoxine, 700 fig. folic acid, 90 pg. biotin, and 11 Mg. vitamin BIT protein content of 47.470 (N X 6.25) and a lysine content of 4.1y0. All lysine values reported were determined by microbiological assay using Leuconostoc mesenteroidea P-60 (8). Day-old New Hampshire chicks were divided into nine lots of six each with due regard to weight and sex. One group received the semipractical basal ration shown in Table I. This ration was found to contain 0.23% lysine by actual analysis. Four lots were fed the basal diet supplemented with 0.28,0.48,0.68, and 0.88% L-lysine.HC1.3 Torula yeast was fed to the remaining four groups at levels of 5, 10,15, and 20y0 of the basal diet. The addition of lysine or protein was accompanied by an equivalent deduction in the protein of the basal diet as derived from corngluten meal and zein. The corn starch of the basal ration was reduced to corre3 Ninety-five per mington, Delaware.
cent
pure
product
obtained
from
E. I. DuPont
and
Co.,
Wil-
416
TSIEN,
Composition Mixture A n-Arginine n-Histidine
JOHNSON
AND
TABLE II of Amino Acid Mixtures Amino acid . HCI . HCI . Hz0
L-Leucine nn-Methionine nn-Phenylalanine nn-Threonine nn-Tryptophan nn-Valine nn-Alanine Dr.-Aspartic acid L-Cystine n-Glutamic acid L-Tyrosine Glycine A
13,521
Mixture B nn-Methionine nn-Tryptophan n-Histidine nL-Phenylalanine Total Mixture B Total Mixture A and Mixture
Used in Rat Studies nag./100g. ration 863 451 1729 1644 468 693 1040 241 1872 577 648 334 2450 660 51
DL-ISOhCiDf?
Total Mixture
LIBlNER
300 40 80 140 B
560 14,081
spond to the nonprotein material introduced by the various levels of To&a yeast. Diets and water were provided ad l&turn for 3 weeks during which time weekly weight changes and total food consumption were recorded. At the completion of the experiment, the animals were sacrificed and the carcasses stored in the freezer for nitrogen analysis. Kjeldahl nitrogen was determined on a representative sample of the ground carcass. Nitrogen retention was aalculated as the difference between the N content of each carcass at completion of the experiment, and the N content of the carcasses from day-old chicks sacrificed and frozen at the beginning of the experiment (average of 20 birds, 10 males and 10 females). The difference in N carcass values per unit of body weight before and after experimentation was likewise calculated. Young rats weighing an average of 60 g. were divided into seven groups with six animals equally allotted to each group on the basis of age, sex, and initial weight. One group was fed a basal purified ration containing the amino acid mixtures A and B (14%) shown in Table II, 5% corn oil,’ 4% salt mixture (9), 74% ’ Mazola.
AVAILABILITY
OF YEAST LYSINE
417
corn starch, 1% corn oil-fat soluble vitamin mix, and 2% sucrose-water soluble vitamin mix.6 The latter two components were so formulated as to provide all the known vitamins (9). Three groups of rats were fed the basal ration to which had been added 0.3,0.6, and 0.9% n-lysine .* The remaining three groups were fed the basal ration supplemented with 7,14, and 21% Torula yeast. In order to maintain the nitrogen of the ration at a constant level, the addition of synthetic lysine or protein in the form of Torula yeast was compensated for by an equivalent reduction in amino acid mixture A. The proportion of amino acid mixture B in the basal ration was kept constant to insure an adequate supply of methionine, tryptophan, histidine, and phenylalanine which tends to be marginal in Torula yeast protein (7). Appropriate reductions in corn starch were made corresponding to the nonprotein contribution of the yeast supplement. All rats were individually housed and given food and water ad lib&m. Weight changes and food consumption were recorded for a period of 3 weeks. In the second week of the preceding experiment, two animals from each group were randomly selected for Iysine absorption studies. Feces were quantitatively collected for a period of 11 days. The fecal pellets could be readily separated by hand from any spilled feed which frequently fell beneath the cages. The absorption of lysine was calculated from data based on the lysine content of the ration and the feces in conjunction with the quantity of food consumed and feces excreted during the period of observation. RESULTS
In Fig. 1 is plotted the growth responseof chicks fed the basal ration (Table I) supplemented with graded levels of synthetic lysine or its equivalent in the form of Torulu yeast as indicated by microbiological analysis. Additional data pertaining to feed efficiency and nitrogen retention are presented in Table III. Up to approximately 0.5 % synthetic or natural lysine there appears to be no significant difference6 in the growth responseelicited by synthetic lysine or yeast providing an equivalent level of lysine. Synthetic lysine in excess of 0.68 % produced no further increment in weight gain; the total level of lysine in such a diet is 0.91% which may be compared with the level of 0.9 % recommended by the National Research Council (10). The superiority of Torula yeast at levels of 15 and 20 % over diets providing comparable levels of synthetic lysine is readily apparent from Fig. 1, and its significance was verified by statistical analysis.E Feed efficiency and nitrogen retention on a per-chick basis closely paralleled the pattern of growth response,with the highest levels of yeast ‘Modified to include vitamin Blz at a level of 4 ag./106 g. ration. (LSignificant differences as used hereafter in this paper will imply a “P” value <0.05 calculated from the analysis of variance. Intertreatment comparisons were made from tables published by May (18).
418
TSIEN,
JOHNSON
t I
AND
I I I
LIENER
I , ,
, ‘
0
0.2 0.4 0.6 0.8 1.0 % L- LYSINE ADDED I I I # I , , , , , J 0 5 10 15 20 25 %TORULA YEAST ADDED FIG. 1. Growth responseof chicks to gradedlevels of L-lysine (0 -0)
or
Torda yeast (*---0) added to basal lysine-deficient diet. The lower abcissa referring to levels of Torula yeast has been adjusted to coincide with the upper
abcissabasedon a lysine content of 4% asdeterminedby microbiologicalanalysis. TABLE III Feed Supplement
E&iency to basal
and Nitrogen ration
None
0.28% L-lysine 0.48yo L-lysine 0.68% L-lysine
Data
in Chick Nitrogen g./ckick
6.3 3.7 2.7 2.2 2.4 4.2 2.6 2.2 2.1
0.60 1.87 2.82 3.85 3.70 1.22 3.02 4.11
Studies retentiona #./kg. body
WA
2.7
19.4
4.91
25.4 40.5 55.5 8.3 21.2 35.1 42.1
between initia1 and final values as described
in the text.
0.88% L-lysine 5% To&a yeast 10% Torula yeast 15% Torula yeast 20% To&a yeast = Difference
Retention Feed efficiency g. feed/g. gain
producing a more efficient utilization of the feed and a greater retention of nitrogen than could be accounted for on the basis of its lysine content. When nitrogen retention is expressed on a per unil body weight basis, however, the retention of nitrogen from To&a yeast was consistently
AVAILABILITY
OF
YEAST
LYSINE
419
less than that from the lysine-supplemented diets, the former averaging about 75 % of the latter. If the lysine of Torula yeast is not completely available as the nitrogen retention data would indicate, then animals receiving 15 % yeast should respond to further additions of synthetic lysine. In a separate experiment it was found that chicks receiving the basal diet plus 15 % To&a yeast gained 109 g. in 3 weeks, whereas a similar lot of chicks receiving an additional supplement of 0.2% n-lysine gained 155 g. This would indicate that superior growth of chicks on high levels of Torula yeast can be attained even though the available lysine provided therein may be suboptimal for growth. Rat Studies The results of the growth study with rats are presented in Fig. 2. At a lysine level of 0.6 % or less, the growth curves of rats receiving synthetic lysine or its equivalent in the form of Toruh yeast were superimposable. The 21% level of Torula yeast, however, produced a significantly greater growth response than one would predict on the basis of its lysine content. In common with the experience of others (11,12), protein efficiency
0 I 0
0.2 0.4 0.6 08 1.0 % L-LYSINE ADDED R I1 II I I I fi I1 5 IO 15 20 25 %TORULA YEAST ADDED
FIG. 2. Growth response of rats to graded levels of L-lysine (O0) or To&a yeast (a- - - -@) added to basal lysine-deficient diet. Adjustments of abcissas were made as described in Fig. 1.
420
TSIEN,
Protein Supplement
Eflciency
to basal
ration
JOHNSON
AND
LIENER
TABLE IV and Lysine Absorption Protein
Data in Rat Studies
e5ciency’
Lysine mg./rot/day
None 0.3% L-lysine 0.6% L-lysine 0.9% L-lysine 7% Torula yeast 14yoTot-da yeast 21o/oTort&la yeast
-0.90 0.15 1.34 1.76 0.13 1.17 2.03
0 43 116 169 33 110 169
0 Grams gain in wt./g. protein consumed. b Milligrams lysine ingested - (mg. lysine excreted basal diet). c (Lysine absorbed/lysine ingested) X 100.
absorbedb
n&/g. body wt./day
%c
0 0.67 1.21 1.67 0.54 1.17 1.39
99.5 98.9 99.9 91.4 90.9 90.2
- mg. lysine excreted on
values (Table IV) were found to parallel growth response, and also showed a significant difference at the highest level of lysine and Torula yeast supplementation in favor of the latter. With reference to the lysine absorption data in Table IV, the same total quantities of lysine were absorbed from both the synthetic and yeast-supplemented diets at essentially equivalent levels of lysine. In proportion to body weight, however, the absorption of lysine from the Torula yeast diets were consistently inferior to the synthetic diets and averaged about 88 % of the latter. In relation to the amount of lysine ingested, the yeast lysine was about 90% as efficiently absorbed as the synthetic amino acid. The fact that the rats receiving the yeast diets absorbed the same total quantity of lysine as those fed the synthetic diets, in spite of the decreased availability of this amino acid from the former, is most likely due to the consumption of greater amounts of food, a logical consequence of enhanced growth. DISCUSSION
Although the primary object of the present study was to evaluate the availability of lysine from Torula yeast, the growth responseswhich were obtained were not solely attributable to this factor. The measurement of lysine availability on the basis of growth alone (or values calculated therefrom such as feed or protein efficiencies), as proposed by Schweigert and Guthneck (5), could not therefore be applied to Toruh yeast. Thus, from the growth curves of Figs. 1 and 2, one would be forced
AVAILABILITY
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LYSINE
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to conclude that at lower levels of Torula yeast the lysine was completely available; whereas at the higher levels of yeast intake the impossible conclusion would be reached that the availability of lysine exceeded 100 %. On the other hand, measurements involving nitrogen retention in chicks and lysine absorption in rats, expressed on the basis of unit body weight, revealed that 75 and 90 % of the lysine of Torulu yeast was available to the chick and rat, respectively. In spite of the fact that the availability of yeast lysine was somewhat less than that of the synthetic product, greater growth responses were obtained with both chicks and rats at levels of Torula yeast ranging from 15 to 20 % of the diet. Since this enhanced growth was observed at levels of yeast calculated to provide lysine sufficient for optimal growth on the purified diets, unrecognized growth factors may well be implicated here. It is also possible that the substitution of yeast protein for the protein of corn-gluten meal and zein (in the chick experiments) or amino acid mixture h (in the rat experiments) may have produced a more favorable balance with respect to amino acids or minerals. The curious finding that the retention of nitrogen per unit body weight of the more rapidly growing chicks fed Torula yeast was less than that of their counterparts receiving synthetic lysine may be due to the preferential deposition of a nonprotein constituent such as fat, an effect similar to that produced by vitamin Blz on the body composition of the rat (13). In addition to Torula yeast being a rich source of the vitamin B complex (14)) these studies further establish the unique value of Torula yeast as a protein supplement and a possible source of unidentified growth factors. However, in view of the adverse physiological effects which high levels of yeast are known to produce, such as liver damage (15)) calciumphosphorus imbalance (16)) and methionine deficiency (17)) further study is warranted before the extensive use of Torula yeast can be recommended.
SUMMARY The availability of lysine from Tom.@ yeast has been measured with chicks and rats on the basis of growth response, feed or protein efficiency, nitrogen retention (chicks), and lysine absorption (rats). Consistently greater growth responses were obtained on lysine-deficient diets supplemented with 15-20 % Torula yeast than could be accounted for on the basis of the lysine so provided. Measurements involving weight gains,
422
TSIEN,
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AND
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therefore, did not prove to be adequate criteria for evaluating lysine availability. On the other hand, nitrogen retention data disclosed that yeast lysine was about 75 % as available to the chick as the synthetic substance. With rats, yeast lysine was about 90 % as efficiently absorbed as the synthetic amino acid. REFERENCES 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17, 18.
BLOCK, R. J., AND MITCHELL, H. H., Nutrition Abstr. and Rev. 16,249 (1946-47). Anonymous, Nutrition Rev. 7, 86 (1949). WHITE, J., “Yeast Technology,” p. 302. Chapman and Hall, London, 1954. KUIKEN, K. A., AND LYMAN, L. M., J. Nutrition 36,359 (1948). SCHWEIGERT, B. S., AND GUTHNECK, B. T., J. Nutrition 49, 277 (1953). GUTHNECK, B. T., AND SCHWEIGERT, B. S., J. Nutrition 49, 289 (1953). INSKEEP, G. C., WILEY, A. J., HOLDERBY, J. M., AND HUGHES, L. P., Ind. Eng. Chem. 43, 1702 (1951). SNELL, E. E., Advances in Protein Chem. 2, 85 (1945). SCHULTZE, M. O., J. Nutrition 41, 103 (1950). National Research Council, “Nutrient Requirements for Poultry.!! Natl. Research Council Publ. 301, 27 (1954). HEGSTED, D. M., AND WORCESTER, J., J. Nutrition 33,685 (1947). SHERWOOD, F. W., AND WELDON, V., J. Nutrition 49, 153 (1953). LING, C. T., AND CHOW, B. F., J. Biol. Chem. 198, 439 (1952). DUNN, C. G., Wallerstein Labs. Communs. 16, 61 (1952). SCHWARZ, K., Ann. N. Y. Acad. 2%. 67, 615 (1954). THAYSEN, A. C., Food 14, 116 (1945). KLOSE, A. A., AND FEVOLD, H. L., J. Nutrition 29, 421 (1945). MAY, J. M., Biometrika 39, 192 (1952).