Determination of Available Amino Acids and Energy in Alfalfa Meal, Feather Meal, and Poultry By-Product Meal by Various Methods1

Determination of Available Amino Acids and Energy in Alfalfa Meal, Feather Meal, and Poultry By-Product Meal by Various Methods 1 Y. HAN and C. M. PARSONS2 Department of Animal Sciences, University of Illinois, Urbana, Illinois 61801 (Received for publication November 14, 1989)

1990 Poultry Science 69:1544-1552 INTRODUCTION

Considerable attention has focused on the precision-fed cockerel assay for determination of digestibility of amino acids (AA) in feedstuffs for poultry (Likuski and Dorrell, 1978; Sibbald, 1979). Because this assay involves quantitative collection of excreta (feces and urine) for 48 h, the procedure actually measures metabolizable AA. However, the term "digestible" or "digestibility" is commonly used because the urinary contribution to excreta AA is small (Bragg et ai, 1969; Terpstra, 1977). Compared wiui growth assays, which are widely accepted as the standard for determining availability of AA, the precisionfed cockerel assay is rapid and inexpensive, and it provides digestibility estimates of several or all AA in feed ingredients. Both growth assays and digestibility or balance assays have shortcomings. In most growth assays, a basal diet deficient in the test AA is supplemented with graded levels of test AA or test feedstuff, and AA availability is

'Supported by Commodity Credit Corporation, USDA, Washington, DC, and by the Illinois Agricultural Experiment Station Project 20-0374. •^To whom reprint requests should be addressed: 322 Mumford Hall, 1301 W. Gregory Drive, Urbana, IL 61801.

calculated from the ratio of the slopes of the respective growth response lines. A problem with this approach is that the growth responses are a function of both basal diet AA and supplemental AA source consumption. Under ad libitum feeding conditions, feed intakes of animals vary among dietary treatments, and animals fed diets supplemented with highly available crystalline test AA usually consume more feed and, thus, more basal diet AA than those fed diets supplemented with the test feedstuff. Partitioning of growth to reflect only that due to supplemental test AA or feedstuff consumption has been suggested as a method of circumventing this potentially confounding problem (Netke and Scott, 1970; Parsons, 1986; Hirakawa and Baker, 1986). The latter procedure has been examined for only a few feedstuffs, and further evaluation is needed. The main criticism of digestibility or balance assays concerns the effects of the hindgut microflora on AA excretion. Consequently, several researchers have proposed the use of cecectomized (CEC) cockerels for determining AA digestibility of feedstuffs (Parsons, 1985, 1986; Johns et al., 1986; Green et al., 1987). Research studies with CEC birds, however, have produced variable results. Parsons (1986) reported that digestibility values of meat meal determined with CEC birds were lower than those determined with

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ABSTRACT Several experiments were conducted to evaluate various methodologies for determining availability of amino acids (AA) and energy in alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM). Digestibility of AA was determined with 48-h precision-fed cockerel assays using conventional (CONV) and cecectomized (CEC) cockerels. Amino acid bioavailability was assessed with chick growth assays. True digestibility values of most AA were significantly lower for CEC than for CONV cockerels. In the chick growth assays, partitioning weight gains to reflect only growth attributable to supplemental crystalline AA or test feedstuff AA consumption often yielded higher bioavailability values than when total weight gains were used for FM and PBPM but lower ones for AM. The magnitude of differences between AA digestibility (cockerels) and AA bioavailability (chicks) values varied among feedstuffs; the largest differences were observed for FM. The TME,j values of AM and PBPM were lower for CEC birds than for CONV birds, whereas TMEn of FM was similar for both types of birds. {Key words: amino acids, energy, alfalfa meal, feather meal, poultry by-product meal, chicken)

AVAILABLE AMINO ACIDS AND ENERGY

MATERIALS AND METHODS

Animals Mature Single Comb White Leghorn cockerels, approximately 45 wk of age, were used in the true digestibility assays. The birds were housed in an environmentally controlled room and kept in individual cages with raised wire floors and subjected to 16 h of light daily. Feed and water were supplied for ad libitum intake before the start of the experiments. Cecectomy was performed according to the procedure of Parsons (1985). The CONV cockerels received a sham operation within a similar time period. All cockerels were given at least 8 wk to recover from surgery prior to being used in experiments. One-wk-old male chicks resulting from the cross of New Hampshire males and Columbian Plymouth Rock females were used in all chick growth assays to estimate bioavailability of Lys, Met and TSAA (Met plus Cys). The chicks were fed a 24% CP corn and soybean meal pretest diet during the 1st wk posthatching. Following an overnight fast, the chicks were weighed, wingbanded, and allotted to dietary treatments as described by Sasse and Baker (1973). Triplicate

groups of five chicks were assigned to each dietary treatment and each assay lasted for 8 or 9 days. All of the chicks were housed in mermostatically controlled starter batteries with raised wire floors in an environmentally regulated room. Feed and water were supplied for ad libitum intake, and light was provided 24 h daily. True Digestibility Assays Three precision-fed cockerel assays were conducted to determine true digestibihty of AA in AM, FM, and PBPM. The assay procedure was that described by Sibbald (1979) with some minor modifications. The number of cockerels on each treatment varied from 6 to 8. Following a 24-h fast, CEC and CONV cockerels were given 30 g of a test feedstuff via crop intubation. A similar number of cockerels of each type were fasted throughout the experimental period to measure endogenous excretion. A plastic tray was placed under each cage and excreta were collected quantitatively for 48 h. The excreta samples were lyophilized, weighed, and ground to pass through a 60-mesh screen. Gross energy, N (Association of Official Analytical Chemists, 1980) and AA content (Spackman et al, 1958) were analyzed on each excreta sample with at least two replicates per sample. Analyses of Met and Cys were performed separately after performic acid oxidation by the method of Moore (1963) with the exception that the excess performic acid was removed by lyophilization after dilution with water. Chick Growth Assays A purified crystalline AA basal diet (Table 1) was employed to determine bioavailability of Lys, Met, and TSAA in most chick growth assays (Baker et al, 1979). Graded levels of the test AA were added to the basal diet containing a suboptimal level of the test AA to produce a reference growth curve. Likewise, increasing levels of test feedstuffs were added to the basal diet at the expense of cornstarch to provide amounts of AA potency that would fall within the boundaries of the reference curve. Lysine bioavailabilities of AM and PBPM were also determined with a Lys-deficient, primarily intact protein diet (Table 2) containing corn, corn gluten meal, and soybean meal to provide a comparison to the purified diet. In the assay for TSAA, the basal diet was devoid of Cys and deficient in Met. This

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conventional (CONV) birds and were also in better agreement with bioavailability values determined by chick growth assays. Raharjo and Farrell (1984a) found little or no difference in AA digestibility between CEC and CONV cockerels, but they found large differences between CONV and ileal-cannulated cockerels (Raharjo and Farrell, 1984b). Johns et al. (1986) and Green et al. (1987) reported that cecectomy yielded lower digestibility values for AA in some feedstuffs but not others. Crissey and Thomas (1987) found that the 48-h excreta collection assay using either CEC or CONV cockerels was not sensitive in detecting reduced AA availability in overcooked soybean meal. The purpose of the present study was 1) to investigate further the effects of cecectomy on AA excretion and AA digestibility values determined by precision-fed cockerel assay and 2) to compare these digestibility values with growth assay bioavailability values calculated by various methods. Alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM), three ingredients varying greatly in composition, were selected for evaluation.

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1546

HAN AND PARSONS TABLE 1. Composition of the crystalline amino acid diet used in the chick growth assays Basal diet

Amino acid mixture

(%)

Amino acid

Cornstarch Amino acid mixture Corn oil Mineral mixture1 Solka floe NaHC0 3 Choline-Cl Vitamin mixture DL-a-tocopheryl acetate (20 mg/kg)

to 100.00 20.49 10.00 5.37 3.00 1.50 .20 .20 +

L-Arg-HCl L-HisHClH 2 0 L-LysHCl3 L-Tyr L-Trp L-Phe L-Met3 L-Cys3 L-Thr L-Leu L-fle L-Val Gly L-Pro L-Glu

Ethoxyquin (125 mg/kg)

+

(%) 1.15 .45 1.15 .45 .15 .50 .35 .35 .65 1.00 .60 .69 .60 .40 12.00

1 Mineral mix provided per kilogram of diet: CaC0 3 , 30 g; Ca 3 (P04) 2 , 28.0 g; K2HPO4, 9.0 g; NaCl, 8.8 g; MgS04-7H 2 0, 3.5 g; MhS0 4 H 2 0, .65 g; ferric citrate, .5 g; ZnC0 3> .1 g; CuS0 4 -5H 2 0, 20.0 mg; H ^ C ^ , 9.0 mg; NaMo0 4 -2H 2 0, 9.0 mg; KI, 40.6 mg; CoS0 4 -7H 2 0, 1.0 mg; Na 2 Se0 3 , .215 mg. ^Vitamin mix provided per kilogram of diet: thiamin-HCl, 20 mg; niacin, 50 mg; riboflavin, 10 mg; Ca-pantothenate, 30 mg; vitamin Bj2> -04 mg; pyridoxine-HCl, 6.0 mg; biotin, .9 mg; folic acid, 4 mg; menadione, 2.0 mg; ascorbic acid, 250.0 mg; retinyl acetate, 5,200 IU; cholecalciferol, 600 IU. Levels of lysine, methionine, and cystine were varied among assays.

procedure was necessary to ensure that all of the bioavailable Cys in FM (very high) would be utilized as a source of TSAA. For determining bioavailability of Met per se, the basal diet was adequate in Cys and deficient in Met. Feather meal was not evaluated in this assay and the lysine bioassay using an intact protein diet because of insufficient quantity of sample. Treatment diets in the growth assays were not maintained as isocaloric in order to evaluate the partitioning procedure (Parsons, 1986) among feedstuffs varying markedly in ME. Statistical Calculations and Analyses True digestibility of AA was calculated according to the method of Sibbald (1979) and TMEn by the method of Parsons et al. (1982). Experimental data were analyzed by analysis of variance procedures (Steel and Torrie, 1980). Statistical Significance of differences in TMEn values and true digestibility values of individual AA between CEC and CONV cockerels were assessed with Student's t test. Slope ratio methodology (Finney, 1978) was used to estimate bioavailability of AA in me chick growth assays. Multiple regression equations were calculated with chick weight gains (grams) as the dependent variable and intake of

TABLE 2. Composition of intact protein basal diet deficient in lysine Ingredient

Amount

(%) Ground corn Cornstarch Com gluten meal (60% CP) Soybean meal (48.5% CP) Corn oil Dicalcium phosphate Calcium carbonate Salt, iodized Ferric citrate Manganese sulfate Zinc carbonate Vitamin premix1 Choline chloride (100%) Selenium premix2 DL-Met L-ArgHCl L-Trp L-Thr

38.14 25.04 22.75 7.25 2.00 2.20 1.00 .40 .06 .05 .01 .10 .10 .01 .03 .65 .09 .12

'Supplied the following amounts per kilogram of diet: vitamin A, 4,400 IU; vitamin D 3 ,1,000 ICU; vitamin E, 11 IU; vitaminBi2, .011 mg; riboflavin,4.4mg; d-pantothenic acid, 10 mg; niacin, 22 mg; menadione sodium bisulfite complex, 2.33 mg. ^Contains 1 mg/g Se from sodium selenite.

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Ingredient

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AVAILABLE AMINO ACIDS AND ENERGY

RESULTS

True Digestibility Assays The composition of AM, FM, and PBPM is given in Table 3. Compared with CEC cockerels, true digestibilities of DM were consistently higher for CONV cockerels; the difference was significant (P<.05) for AM (Table 4). The TMEQ values of AM and PBPM were significantly (P<05) greater for CONV than for CEC cockerels. Apparent N retention was similar between types of birds within feedstuff. True digestibility of more than half of the measured AA in the three feedstuffs was significantly lower (P<.05) for CEC than for CONV cockerels. Digestibility of Cys was relatively low in all of the feedstuffs. Comparing average digestibility of all the AA, the differences between CEC and CONV birds were largest for AM (67.6% versus 79.4%), smallest for FM (69.2% versus 74.5%), and intermediate for PBPM (78.2% versus 85.9%). Chick Growth Assays Body weight gains of chicks responded linearly to supplementation of the AA-deficient basal diets with crystalline AA or the feedstuffs in all assays (Tables 5 to 8) except for FM supplementation in the first Lys assay (Table 5). Partitioned weight gains responded similarly. With multiple linear regression analysis and slope ratio methodology applied to total chick weight gains, Lys bioavailability was calculated to be 88.2% for AM, 69.9% for PBPM, and only 21.0% for FM when a purified diet was used (Table 5). Bioavailability values of Lys com-

TABLE 3. Composition of feed ingredients (g/100 g)1

Component Moisture Crude protein Ether extract Ash Amino acids Asp Thr Ser Glu Pro Gly Ala Cys Val Met He Leu

Tyr Phe Lys Arg Total

Alfalfa meal

Feather meal

Poultry by-product meal

8.90 16.00 1.78 10.53

6.22 86.25 5.81 2.79

9.14 58.10 12.26 16.50

2.08 .74 .75 1.65 1.23 .83 .87 .23 .88 .28 .67 1.24 .43 .78 .78 .72 14.16

5.86 4.03 9.36 10.18 8.35 6.00 3.94 4.52 6.64 .40 4.26 7.18 2.47 4.17 2.02 6.14 85.52

5.53 2.33 2.99 7.49 4.79 6.73 4.04 1.29 3.12 1.10 2.11 4.40 1.85 2.47 2.94 4.40 57.58

Values are expressed on an air-dry basis.

puted from partitioned gains were higher than those from total gains for FM and PBPM, whereas bioavailability was lower for AM. However, these differences were not significant (P>.05). Standard errors of the bioavailability estimates were large for AM and FM, and partitioning of weight gains did not reduce them. Consequently, the regression coefficients and slope ratio estimates for AM were not significantly different from 100% and those for FM were not significantly different from zero. When an intact protein diet was fed, bioavailability of Lys computed from total weight gains was 92.6% for AM and 62.9% for PBPM (Table 6). When computed from partitioned weight gains, the bioavailability values were 65.9% for AM and 73.9% for PBPM and were significantly (P<.05) different than the respective values calculated from total weight gains. The precision of the bioavailability estimates was much better than in the previous Lys assay. Bioavailability of TSAA based on total weight gains was estimated to be 79.4% for AM, 63.3% for FM, and 60.3% for PBPM (Table 7). Partitioning the weight gains had no significant effect on bioavailability values. Bioavailability of Met based on total weight gains and determined with a Met-deficient, Cys-

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supplemental test crystalline AA and feedstuff AA (milligrams) as the independent variables. In the TSAA assay, intake of sulfur (millimole) from Met and Cys was used as the independent variable in computing the multiple regression to account for molecular weight differences between the AA. The bioavailability of AA in each feedstuff was then estimated by the ratio of the slope of its regression line to that of the crystalline test AA, which was assumed to be 100% bioavailable. Bioavailability was computed by an additional method wherein weight gains of chicks were partitioned to reflect only gain attributable to intake of supplemental AA in crystalline form or to supplemental AA from the test ingredient (Parsons, 1986).

1548

HAN AND PARSONS

adequate purified diet was 73.4% for AM and 57.4% for PBPM (Table 8). Estimates of Met bioavailability were 64.2% for AM and 70.1% for PBPM when calculated from partitioned weight gains. From the results of the TSAA and the specific Met growth assays, the bioavailability of Cys was calculated by difference. These calculations yielded bioavailable Cys values of 84.3 and 62.8% for AM and PBPM when calculated from total weight gains, and 74.6 and 51.8% when based on partitioned weight gains, respectively.

In general, the composition data for AM, FM, and PBPM data agreed with those listed by the National Research Council (NRC, 1984) except for a few AA in FM and PBPM. These differences are probably due to variation in the raw materials or processing conditions. The total of the AA in AM was lower than the CP value; however, the levels of analyzed AA were in general agreement with the NRC

TABLE 4. Influence of cecectomy on utilization of alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM) in the precision-fed cockerel assay AM

PBPM

FM

Measurement

CONV1

CEC1

SED1 CONV

CEC

SED

CONV

CEC

SED

Number of observations True digestibility of DM, % N retained (g), apparent TMEn, kcal/g DM True digestibility of amino acids, % Asp Thr Ser Glu Pro Ala Cys Val Met He Leu Tyr Phe Lys Arg Mean

8 21.5* -1.41* 1.193*

7 11.7b -1.36* .904b

8 3.8 44.3* .25 -.55* .098 3.219*

8 41.4* -.62* 3.131*

3.3 21 .065

7 57.0* .01* 3.390*

6 49.6* .13* 2.863b

4.1 .24 .082

81.6* 76.9* 77.4* 81.2* 86.4* 78.8* 61.5* 80.5* 75.1* 83.1* 83.9* 81.1* 84.5* 73.4* 85.5* 79.4

73.5b 61.5b 59.9 b 71.4 b 77.8b 69.4b 30.5b 70.2b 69.7* 76.7* 79.7* 58.8b 72.9b 63.6* 77.7* 67.6

1.6 2.8 3.4 2.6 2.3 2.5 9.8 2.4 4.1 3.0 4.2 5.3 3.3 4.6 4.5

82.5* 84.9* 85.8* 86.2* 87.8* 87.3* 81.3* 82.8* 90.1* 85.8* 87.0* 87.1* 87.4* 83.1* 88.7* 85.9

58.7b 75.3 b 74.3 b 76.3 b 78.4b 80.8b 68.5b 81.3* 85.9b 81.2b 84.0* 80.3* 84.6* 75.3 b 84.9b 78.2

59.2* 69.8* 78.4* 68.7* 71.6* 80.8* 60.4* 84.5* 62.3* 87.5* 82.1* 76.8* 85.6* 68.9* 81.3* 74.5

43.2 b 66.5* 70.5b 61.5b 63.7b 78.0 b 47.2 b 83.0* 54.8 b 86.4* 79.6b 75.5* 83.9* 66.7* 77.7b 69.2

2.3 2.8 1.3 1.6 1.7 1.3 3.3 1.0 3.2 .9 1.2 1.5 .9 2.0 1.4

2.7 2.2 22 2.0 1.6 1.5 2.2 3.3 1.5 2.1 1.9 3.4 1.9 22 1.6

*-bWithin feedstuffs, means in the same row with no common superscripts are significantly different (P<05). CONV = conventional; CEC = cecectomized; SED = standard error of the difference calculated a s ^ s /N when sample sizes were equal and calculated based on weighted average of sample variances when sample sizes were unequal (Steel and Torrie, 1980).

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DISCUSSION

(1984) and the values reported by Sibbald (1986) for AM of similar CP content. One uncertainty of an excreta collection digestibility assay is the unknown contribution of hindgut microbial AA metabolism to AA content of excreta. The present study showed that the influence of cecectomy on AA digestibility values was substantial; CEC birds yielded significandy lower digestibility values than CONV birds in most cases. There was no difference observed in N balance between CEC and CONV cockerels given the same feedstuff even though digestibilities of AA differed. These results suggest substantial deamination of AA to ammonia, which was probably subsequently absorbed from the ceca and then excreted via the urine (Mortensen, 1984). Significant differences in TMEn values between bird types were observed for AM and PBPM but not for FM. Kessler and Thomas (1981) reported mat TME values for soybean meal were higher for CONV than for CEC cockerels, and Parsons (1986) observed similar results for meat meal. The higher TMEn

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AVAILABLE AMINO ACIDS AND ENERGY

TABLE 5. Determination of lysine bioavailability in alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM) using a slope ratio chick bioassay with a purified diet1 Bioavaliability3

Weight gain Treatment Basal (B) 4 B + .15% L-Lys B + .30% L-Lys B + 5% AM B + 10% AM B + 15% AM B + 5% FM B + 10% FM B + 3% PBPM B + 7% PBPM B + 11% PBPM Pooled SE 1

Total 31.8 55.5 81.4 37.8 41.5 45.1 31.6 38.9 40.5 48.6 60.1 2.8

2

Partitioned -2.3 14.8 31.4 2.7 3.9 5.1 -1.0 5.8 5.2 12.4 21.5 2.4

Feed intake 74.7 93.6 120.1 77.5 84.7 91.5 70.4 71.9 78.3 81.0 87.7 3.6

Gainrfeed

Mean

(g:g) .425 .594 .682 .487 .485 .494

— (% of total) — 100

SE

882 (66.6

18.8 27.3)

.449 .542

21.6 (38.0

14.7 20.7)

.518 .603 .685 .026

69.9 (85.8

7.1 10.5)

Means of triplicate groups of five male chicks from 8 to 16 days postnatching. Weight gains partitioned to reflect only gain due to supplemental crystalline amino acid or feedstuff amino acid intake. The equation derived from the first three treatments by regressing total gain (Y; grams) on total Lys intake (X; milligrams) was Y = 7.85 + .08773X, r 2 = .93. Calculated from slope ratio analysis for multiple regression of total or partitioned weight gain (grams) on supplemental crystalline lysine or feedstuff lysine intake (milligrams). The regression equations were Y = 33.09 + .1367 (L-Lys) + .1205 (AM) + .02949 (FM) + .09560 (PBPM), R 2 = .92 and Y = -.46 + .08985 (L-Lys) + .05988 (AM) + .03414 (FM) + .07713 (PBPM), R 2 = .85, respectively. Values in parentheses calculated using partitioned gains. TJasal diet was a purified crystalline amino acid diet (Table 1) adequate in all amino acids except Lys (.4% L-Lys). 2

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estimates from CONV cockerels suggest sig- crystalline) is usually greater than that of the nificant production of volatile fatty acids by AA from die test feedstuff, differences in feed bacteria in the ceca (Annison et ah, 1968). intake among treatments could influence bioThus, CEC birds may underestimate the availability values. As described by Parsons amount of energy utilized from feedstuffs by (1986), partitioning results in AA bioavailabilCONV birds. The fate of the volatile fatty ity being based only on the growth attributable acids produced in the ceca and the amount of to the supplemental test AA or feedstuff AA useful energy derived from microbial metabo- consumed. Dietary energy was lowered marlism are not well understood (Mortensen, kedly when AM was added to the basal diet in place of cornstarch. The reduced dietary 1984). Bioavailability values for AA in AM were energy combined with only a modest proporconsistently higher when computed from total tion of the total dietary test AA being supplied weight gains than when computed from parti- by AM resulted in markedly increased intake tioned weight gains. However, partitioning of the basal AA and, thus, yielded high AA generally had the opposite effect with FM and bioavailability values. Hence, partitioning PBPM. The latter agrees with previous obser- reduced bioavailability values for AM. Unlike vations for meat meal (Parsons, 1986) and for AM, bom FM and PBPM have caloric values a mixture of corn gluten meal, soybean meal, (TME,,) similar to the dietary cornstarch that and meat and bone meal (Hirakawa and Baker, they replaced. The increase in bioavailability 1986). The partitioning procedure is an attempt from partitioning for these feedstuffs, when to adjust for differences in ad libitum feed occurring, was probably due to the fact mat intake among treatments (Netke and Scott, chicks given diets supplemented with FM or 1970). Because chick growth is a function of PBPM consumed less feed, and thus, less of consumption of both basal diet AA and the highly available basal AA, than did chicks supplemental AA source, and the bioavailabil- given the reference diets. As mentioned earlier, ity of the AA in the basal diet (often the energy content of the diets used herein was

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HAN AND PARSONS TABLE 6. Determination of lysine bioavailability in alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM) using a slope ratio chick bioassay with an intact protein diet1 Weight gain 2

Gain: feed (g:g) .406 .509 .592

— (% of total) 100

.433 .446 .481 .463 .520 .570

92.6 (65.9

5.3 4.9)

62.9 (73.9

2.6 2.5)

Total

Partitioned'

Basal (B) 4 B + .15% L-Lys B + .30% L-Lys B + 7% AM B + 14% AM B + 21% AM B + 4% PBPM B + 8% PBPM B + 12% PBPM

48.7 73.3 107.0 55.4 65.7 74.1 59.7 72.5 83.3 2.2

.0 14.9 33.7

120.2 143.8 180.7

3.5 5.9 11.6 7.4 16.0 24.0

127.9 147.5 154.2

(g/chick)

128.9 139.3 146.3 4.4

Mean

SE

.005

Means of triplicate groups of five male chicks from 8 to 17 days posthatching. Weight gains partitioned to reflect only gain due to supplemental crystalline amino acid or feedstuff amino acid intake. Only Treatment 1 was used to partition weight gain since the basal diet contained intact protein (Parsons, 1986). Average gain (grams) per milligram Lys consumed from the basal diet was .073752. Calculated from slope ratio analysis for multiple regression of total or partitioned weight gain (grams) on supplemental crystalline lysine or feedstuff lysine intake (milligrams). The regression equations were Y = 49.12 + .1077 (L-Lys) + .0998 (AM) + .06781 (PBPM), R 2 = .98 and Y = .49 + .06199 (L-Lys) + .04084 (AM) + .0458 (PBPM), R 2 = .99 for total and partitioned weight gains, respectively. Values in parentheses calculated using partitioned gains. 4 Basal diet was the intact protein diet as presented in Table 2.

TABLE 7. Determination of bioavailability of total sulfur amino acids (TSAA) in alfalfa meal (AM), feather meal (FM), and poultry by-product meal (PBPM) using a slope ratio chick bioassay with a purified diet1 Weight gain Treatment

Total

Partitioned2

Feed intake

(g/chick) Basal (B) 4 B + .10% TSAA5 B + .20% TSAA5 B + 7% AM B + 14% AM B + 21% AM

11.1 44.7 69.7 22.1 29.3 33.2

B B B B B B

28.3 39.4 51.5 32.6 44.1 59.1 2.5

+ + + + + +

1.5% FM 3.0% FM 4.5% FM 4% PBPM 8% PBPM 12% PBPM

Pooled SE

-2.1 14.9 33.7 3.5 5.9 11.6 8.6 14.8 25.0 10.8 19.1 29.4 1.7

58.3 95.5 120.0 78.1 83.5 86.5 78.5 93.7 99.2 85.0 94.9 109.4 3.8

Gain: feed

Bioavailability3 Mean

SE

(g:g) .190 .467 .581

— (% of total) — 100

.283 .350 .382

79.4 (69.9

12.4 12.6)

.358 .420 .518 .385 .464 .540

63.3 (63.6

5.0 5.1)

60.3 (60.2

4.1 4.1)

.019

Means of triplicate groups of five male chicks from 8 to 17 days posthatching. 2 Weight gains partitioned to reflect only gain due to supplemental crystalline amino acid or feedstuff amino acid intake. The equation derivedfromthefirstthree treatments by regressing total gain (Y; grams) on total intake of sulfur from TSAA (X; millimole) was Y = -4.05 + 22.23X, r 2 = .98. Calculated from slope ratio analysis for multiple regression of total or partitioned weight gain (grams) on intake of sulfur (millimole) from supplemental crystalline TSAA or feedstuff TSAA. The regression equations were Y = 17.78 + 30.05 (TSAA) + 23.86 (AM) + 19.01 (FM) +18.12 (PBPM), R 2 = .93 and Y = 1.36 + 21.17 (TSAA) + 14.80 (AM) + 13.47 (FM) + 12.74 (PBPM), R = .93 for total and partitioned weight gains, respectively. Values in parentheses calculated using partitioned gains. 4 Basal diet was a purified diet deficient in TSAA (.2% DL-Met, 0% L-Cys). Supplied as equal amounts of DL-Met and L-Cys.

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Treatment

Pooled SE

Bioavailability3

Feed intake

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AVAILABLE AMINO ACIDS AND ENERGY

The present study also illustrates that growth assays have definite limitations for evaluation of low energy, moderate protein ingredients such as AM or ingredients with very poor AA balance such as FM. Bioavaila-

bility estimates for AM were less precise than those for PBPM in all assays, regardless of computation method. The Lys bioavailability value for FM was very low and very imprecise. Smith (1968) reported similar problems when attempting to determine Lys availability in FM by chick growth assay. The magnitude of difference between AA digestibility and AA bioavailability values varied among feedstuffs. Because only a few feedstuffs were investigated in the present study, a valid correlation analysis could not be conducted. Digestibility values determined with CEC cockerels agreed reasonably well with chick bioavailability values for Lys and Met in AM (partitioned) and Lys and Cys in PBPM, whereas digestibility values from CONV cockerels were somewhat higher. Digestibility of TSAA in FM for CONV birds was similar to bioavailability for chicks. Alternatively, digestibility values did not agree with bioavailability values in other cases; namely, digestibilities of Lys in FM and Met

TABLE 8. Determination of bioavailability of methionine in alfalfa meal (AM) and poultry by-product meal (PBPM) using a slope ratio chick bioassay with a purified diet Weight gain Treatment

Total

Partitioned2

Basal (B) 4 B + .05% L-Met B + .10% L-Met B + 7% AM B + 14% AM B + 21% AM B + 5% PBPM B + 10% PBPM B + 15% PBPM

18.1 39.1 72.6

-.6 12.6 32.7 2.4 5.4 7.0 3.3 16.2 28.3 1.5

Gain: feed

Feed intake -

Pooled SE Bioavailability of Cys Alfalfa meal Poultry by-product meal x

21.9 27.3 33.0 24.3 40.4 55.9 2.6

62.5 84.3 122.5 64.9 71.4 83.2 68.8 78.1 87.9 3.9

(g:g) .290 .463 .592 .337 .380 .396 .353 .516 .636 .013

Bioavaiilability3 Mean

SE

— (% of total) — 100

73.4 (64.2

8.3 10.0)

57.4 (70.1

2.9 3.6)

84.3 (74.6) 62.8 (51.8)

Means of triplicate groups of five male chicks from 8 to 17 days posthatching. Weight gains partitioned to reflect only gain due to supplemental crystalline amino acid or feedstuff amino acid intake. The equation derived from the first three treatments by regressing total gain (Y; grams) on Met intake (X; milligrams) was Y = -3.38 + .2719X, r 2 = .99. Calculated from slope ratio analysis for multiple regression of total or partitioned weight gain (grams) on supplemental crystalline methionine or feedstuff methionine intake (milligrams). The regression equations were Y = 17.31 + .4596 (Met) + .3373 (AM) + .2640 (PBPM), R 2 = .97 and Y = -.82 + .2793 (Met) + .1792 (AM) + .1957 (PBPM), R 2 = .97, for total and partitioned weight gains, respectively. 4 Basal diet was a purified diet deficient only in Met (.13% DL-Met, .30% L-Cys). Calculated by difference using the TSAA bioavailability values in Table 7 and the Met bioavailability values presented in this table. Values in parentheses calculated from partitioned weight gains.

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intentionally not kept isocaloric so that the partitioning procedure could be evaluated. The results indicated that the partitioning procedure may represent a more defensible method for determining AA bioavailability, particularly when energy content of diets varies markedly or feed intakes vary greatly among treatments or both. In most cases, bioavailability estimates from regressions based on partitioned gains are less precise and have slightly larger standard errors than those based on total weight gains. When feasible, it may be better to design bioavailable AA assays so that ME,, content is maintained constant across all dietary treatments to minimize differences in feed intake. In addition, the isocaloric diets could be pair-fed to equalize intake of energy for all treatments.

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REFERENCES Annison, E. F., K. J. Hill, and R. Kenworthy, 1968. Volatile fatty acids in the digestive tract of the fowl. Br. J. Nutr. 22:207-216. Association of Official Analytical Chemists, 1980. Official Methods of Analysis. 13th ed. Assoc. Off. Anal. Chem., Washington, DC. Baker, D. H., R. C. Blitenthal, K. P. Boebel, G. L. Czarnecki, L. L. Southern, and G. M. Willis, 1981. Protein-amino acid evaluation of steam-processed feather meal. Poultry Sci. 60:1865-1872. Baker, D. H., K. R. Robbins, and J. S. Buck, 1979. Modification of the level of histidine and sodium bicarbonate in the Illinois crystalline amino acid diet. Poultry Sci. 58: 749-750. Bragg, D. B., C. A. Ivy, and E. L. Stephenson, 1969. Methods for determining amino acid availability of feeds. Poultry Sci. 48:2135-2137. Crissey, S. D., and O. P. Thomas, 1987. Comparison of the sensitivities of growth and digestibility studies using intact, cecectomized, and cannulated roosters. Poultry Sci. 66:866-874. Finney, D. J., 1978. Statistical Method of Biological Assay. 3rd ed. Charles Griffin and Co. Ltd., High Wycombe, Bucks, England. Green, S., S. L. Bertrand, M.J.C. Duron, and R. Maillard, 1987. Digestibilities of amino acids in soyabean, sunflower and groundnut meals, determined with intact and caecectomised cockerels. Br. Poult Sci. 28: 643-652. Hirakawa, D. A., and D. H. Baker, 1986. Assessment of lysine bioavailability of an intact protein mixture: comparison of chick growth and precision-fed rooster assay. Nutr. Res. 6:815-826. Johns, D. C, C. K. Low, J. R. Sedcole, and K.A.C. James, 1986. Determination of amino acid digestibility using caecectomised and intact adult cockerels. Br. Poult.

Sci. 27:451-461. Kessler, J. W., and O. P. Thomas, 1981. The effect of cecectomy and extension of the collection period on the true metabolizable energy values of soybean meal, feather meal, fish meal, and blood meal. Poultry Sci. 60:2639-2647. Likuski, HJA., and H. G. DorrelL 1978. A bioassay for rapid determination of amino acid availability values. Poultry Sci. 57:1658-1660. Mortensen, A., 1984. Importance of microbial nitrogen metabolism in the ceca of birds. Pages 273-278 in: Current Perspectives in Microbial Ecology. Third International Symposium of Microbiology and Ecology. American Society of Microbiology, Washington, DC. Moore, S., 1963. On the determination of cystine as cysteic acid. J. Biol. Chem. 238:235-237. National Research Council, 1984. Nutrient Requirements of Poultry. Nutrient Requirements of Domestic Animals. 8th rev. ed. Natl. Acad. Sci., Washington, DC. Netke, S. P., and H. M. Scott, 1970. Estimates on the availability of amino acids in soybean oil meal as determined by chick growth assay: methodology as applied to lysine. J. Nutr. 100:281-288. Parsons, C. M., 1985. Influence of caecectomy on digestibility of amino acids by roosters fed distillers' dried grains with solubles. J. Agric. Sci. Camb. 104: 469-472. Parsons, C. M, 1986. Determination of digestible and available amino acids in meat meal using conventional and caecectomized cockerels or chick growth assays. Br. J. Nutr. 56:227-240. Parsons, C. M., L. M. Potter, and B. A. Bliss, 1982. True metabolizable energy corrected to nitrogen equilibrium. Poultry Sci. 61:2241-2246. Raharjo, Y. C , and D. J. Farrell, 1984a. Effects of caecectomy and dietary antibiotics on the digestibility of dry matter and amino acids in poultry feeds determined by excreta analysis. Aust. J. Exp. Agric. Anim. Husb. 24: 516-521. Raharjo, Y. C , and D. J. FarrelL 1984b. A new biological method for determining amino acid digestibility in poultry feedstuffs using a simple cannula, and the influence of dietary fibre on endogenous amino acid output. Anim. Feed Sci. Technol. 12:29-45. Sasse, C. E., and D. H. Baker, 1973. Availability of sulfur amino acids in com and corn gluten meal for growing chicks. J. Anim. Sci. 37:1351-1355. Sibbald, I. R., 1979. A bioassay for available amino acids and true metabolizable energy in feedstuffs. Poultry Sci. 58:668-673. Sibbald, I. R., 1986. The T.M.E. system of feed evaluation: methodology, feed composition data and bibliography. Technical Bulletin 1986-4E, Agric. Canada, Ottawa, Ontario, Canada. Smith, R. E., 1968. Assessment of the availability of amino acids in fish meal, soybean meal and feather meal by chick growth assay. Poultry Sci. 47:1624-1630. Spackman, D. H., W. H. Stein, and S. Moore, 1958. Automatic recording apparatus for use in the chromatography of amino acids. Anal. Chem. 30:1190-1206. Steel, R.G.D., and J. H. Tome, 1980. Principles and Procedures of Statistics. A Biomedical Approach. 2nd ed. McGraw-Hill Book Co., Inc., New York, NY. Terpstra, K., 1977. Determination of the digestibility of protein and amino acids in poultry feed. Pages 1-8 in: Proceedings of Fifth International Symposium on Amino Acids. Budapest, Hungary.

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in PBPM were higher and digestibility of Cys in AM was lower than chick bioavailability values. Which of these values is most accurate is unknown. Possible explanations for the differences between assay type include negative or positive effects of the test protein AA pattern on utilization of the fast-limiting AA in the growth assays (Hirakawa and Baker, 1986) and intestinal absorption of AA derivatives that were not subsequently bioavailable for protein synthesis in the digestibility assays, such as lanthionine and lysinoalanine in FM (Baker et ah, 1981). Although the growth assay is usually considered the standard and most accurate method for determination of AA availability, the precision-fed cockerel digestibility assay with CEC birds has many advantages over the former and appears to be a better assay for routine use. However, the reason for the substantial differences between cockerel digestibility and chick bioavailability values seen in some cases deserves further investigation.