1. Introduction
According to physiological needs, absorbed amino acids (AA) should be available for tissue protein synthesis in time and in a ratio that matches the AA requirement of individual tissues. Finally, recommended optimal dietary AA ratios (ideal protein concepts) summarize these tissue needs for the whole animal. However, both due to variation of individual tissue growth depending on age or performance and the ratio between AA needs for maintenance and performance, the optimal dietary AA ratio cannot be expected as constant. A maximized correspondence between dietary supply and physiological requirements is the tool to improve metabolic efficacy within the physiological possibilities and sustainability of nutrient conversion in systems for food producing animals.
However, both improved AA requirement data and procedures to evaluate the feed potential are needed. Current focus on ileal digestible AA as a tool to describe the feed potential does not take into account that on average, 40 percent of the absorbed AA could be catabolized by the enterocytes in the gut [
1]. Possibilities and limitations of different procedures for AA requirement studies are discussed elsewhere [
2,
3,
4,
5,
6]. A controversy exists according to the procedures, which are mostly adapted to the physiological conditions in the animal. Currently, such procedures utilize dose response studies making use of graded AA supplementation [
7,
8] or derive optimal ratios from AA response due to individual AA deletion from an AA complete diet [
9]. According to Baker [
8], diets are formulated to meet the recommendations of the National Research Council (NRC) [
10], except for the amino acid under study. Estimates of the AA requirement are conducted in terms of “subjective” and “objective” estimates of the required AA concentration in the diet. The “subjective estimate” is concluded from broken-line analysis only. Deriving the first point at which the quadratic response curve intersects the plateau delivered from broken-line analysis provides the “objective estimate”. The latter is utilized for further conclusion of ideal dietary AA ratios.
In addition, Wang and Fuller [
9] have developed another approach based on a created AA complete diet near to the actual assumptions for an optimal dietary AA ratio. This complete diet contained crystalline AA supplementations for each of the AAs under study to supply 100% of the assumed requirement level. In the next step, the AA under study was individually deleted from the complete diet. Both responses were measured, due to the complete and the deleted diet, respectively. The observed slope of the response criteria between these diets was utilized for conclusion of optimal dietary ratios between individual AAs. The principle of this approach was utilized in our procedure, but the effect of individual AA deletion was directly measured by AA efficiency from modelling of observed N balance data in growing chicken [
5]. The aim of present experiments was to apply observed AA efficiency data of lysine, threonine, tryptophan, arginine, isoleucine and valine for conclusions about optimal AA ratios for these AAs in diets of growing chicken.
3. Results and Discussion
Results of the conducted N balance experiments are summarized in
Table 5. From these data, both dietary protein quality (
b) and AA efficiency (
bc−1) data were derived (
Table 6,
Table 7).
Generally, in nearly all experimental periods, lower N balance data were obtained with AA diluted diets as compared to the balanced CD, resulting from both reduced supply of individual AA and total N intakes (
Table 5). A more specific evaluation of the responses due to individual AA dilution was obtained by applying the model parameter,
b. If the limiting position of the AA under study was valid, the parameter,
b, declined significantly. In contrast, when the optimal dietary supply exceeded the yielded response on dietary protein quality due to individual AA dilution was only marginal.
During the starter period in Experiment I, the dilution of LYS, THR and ARG led to significant impairment of the dietary protein quality (
b) and indicated a valid limiting position of these AAs (
Table 6). Other diets yielded no significant effect on
b. On the other hand, following diets TRP, ILE and VAL, the obtained AA efficiency data (
bc−1) were significantly improved. This observation could indicate a minor oversupply of these AAs in the control diet (CD),
i.e., the diluted diets with considerably lower contents of TRP, ILE and VAL yielded equal protein quality as compared to CD. To overcome the excessive supply of individual AAs, Experiment II utilized an elevated lysine concentration in the feed protein (
Table 4). In consequence, due to the dilution diet, TRP also yielded a significantly impaired protein quality (
b). In spite of the indicated limiting position in AA diluted diets, TRP and ARG, the efficiency of both AA were not improved (p > 0.05). It is possible that TRP and ARG act as co-limiting AAs in this CD, because no impact on efficiency (
bc−1) of TRP and ARG was observed. Furthermore, this assumption is supported by significantly lower LYS efficiency in the LYS diluted diet. It cannot be excluded that changes in diet composition due to variable batches and proportions of main feed ingredients (
Table 3), different dietary AA concentrations (
c) with possibly varying AA efficiencies or the slightly lower total N contents in diluted diets (
Table 6) were responsible for this unexpected observation. According to the results of Experiments I and II, diets, ILE and VAL, demonstrated that the concentrations of these BCAA were still too high to achieve the ensured limiting position. Consequently, a higher dilution of Ile and Val down to 70% of CD, as realized in Experiment III, did overcome this problem. The observed response of
b due to diets, ILE and VAL, in Experiment III underlines that both of the AAs achieved their limiting position.
Table 5.
Summarized data of the N balance experiments (mean ± SEM *).
Table 5.
Summarized data of the N balance experiments (mean ± SEM *).
Exp. | Diet | Starter period (11d–21d) | Grower period (25d–35d) |
---|
BW (g) | NI (mg /BWkg0.67/d) | ND ** (mg /BWkg0.67/d) | BW (g) | NI (mg /BWkg0.67/d) | ND** (mg /BWkg0.67/d) |
---|
I | CD | 554 a ± 44 | 3,384 a ± 69 | 2,315 a ± 50 | 1,332 ± 112 | 2,970 a ± 67 | 2,010 a ± 73 |
LYS | 400 b ± 45 | 2,939 b ± 96 | 1,893 b ± 46 | 1,462 ± 52 | 2,633 b ± 14 | 1,707 b ± 29 |
THR | 518 a ± 55 | 3,160 a ± 102 | 2,114 b ± 51 | 1,370 ± 61 | 2,590 b ± 43 | 1,758 b ± 23 |
TRP | 528 a ± 51 | 3,399 a ± 138 | 2,297 a ± 82 | 1,355 ± 135 | 2,743 b ± 18 | 1,957 a ± 47 |
ARG | 575 a ± 37 | 3,137 b ± 62 | 2,077 b ± 65 | 1,147 ± 82 | 2,557 b ± 70 | 1,753 b ± 45 |
ILE | 597 a ± 39 | 3,122 b ± 64 | 2,238 a ± 52 | 1,382 ± 58 | 2,776 b ± 19 | 1,929 a ± 30 |
VAL | 596 a ± 41 | 3,121 b ± 62 | 2,197 a ± 63 | 1,416 ± 71 | 2,582 b ± 43 | 1,758 b ± 32 |
II | CD | 624 ± 53 | 3,711 a ± 60 | 2,500 a ± 32 | 1,880 ± 83 | 3,171 ± 41 | 2,063 a ± 20 |
LYS | 625 ± 46 | 3,462 b ± 46 | 2,027 b ± 29 | 1,758 ± 76 | 3,112 ± 20 | 1,869 b ± 30 |
THR | 688 ± 49 | 3,513 b ± 55 | 2,317 b ± 43 | 1,759 ± 83 | 3,191 ± 30 | 1,940 b ± 25 |
TRP | 640 ± 50 | 3,367 b ± 106 | 2,128 b ± 60 | 1,847 ± 86 | 3,131 ± 33 | 1,960 b ± 33 |
ARG | 673 ± 49 | 3,470 b ± 53 | 2,170 b ± 43 | 1,777 ± 74 | 3,032 ± 63 | 1,842 b ± 35 |
ILE | 672 ± 52 | 3,456 b ± 56 | 2,374 b ± 46 | 1,804 ± 98 | 3,162 ± 37 | 1,906 b ± 22 |
VAL | 674 ± 50 | 3,462 b ± 95 | 2,385 a ± 85 | 1,840 ± 82 | 3,149 ± 27 | 1,882 b ± 36 |
III | CD | 538 a ± 38 | 4,025 a ± 31 | 2,703 a ± 23 | 1,570 a ± 57 | 3,565 a ± 45 | 2,182 a ± 44 |
LYS | 380 b ± 32 | 3,506 b ± 127 | 2,201 b ± 95 | 1,475 a ± 60 | 3,452 a ± 86 | 1,912 b ± 62 |
THR | 468 a ± 41 | 3,771 b ± 83 | 2,503 b ± 66 | 1,469 a ± 52 | 3,449 a ± 68 | 2,037 b ± 45 |
ILE | 441 a ± 37 | 3,752 b ± 88 | 2,410 b ± 52 | 1,291 b ± 31 | 3,328 b ± 85 | 1,872 b ± 59 |
VAL | 387 b ± 34 | 3,459 b ± 126 | 2,361 b ± 77 | 1,309 b ± 71 | 2,715 b ± 167 | 1,562 b ± 134 |
Table 6.
Model parameters as derived from N balance studies with control (CD) and AA diluted (DD) diets in the starter period (mean ± SEM *).
Table 6.
Model parameters as derived from N balance studies with control (CD) and AA diluted (DD) diets in the starter period (mean ± SEM *).
Exp. | Diet | N-content (g/kg) | c (g/16g N) | b (b·106) | bc−1 |
---|
CD | DD | CD | DD |
---|
I | CD | 36.66 | | | 314 a ± 6 | | |
LYS | 33.93 | 5.47 | 4.73 | 268 b ± 3 | 57 ± 1 | 57 ± 1 |
THR | 34.22 | 3.55 | 3.04 | 292 b ± 7 | 88 B ± 2 | 96 A ± 2 |
TRP | 34.34 | 0.99 | 0.85 | 310 a ± 10 | 315 B ± 6 | 366 A ± 12 |
ARG | 33.56 | 5.72 | 5.00 | 287 b ± 8 | 55 ± 1 | 57 ± 2 |
ILE | 34.23 | 3.67 | 3.14 | 322 a ± 6 | 85 B ± 2 | 103 A ± 2 |
VAL | 34.17 | 4.46 | 3.83 | 313 a ± 8 | 70 B ± 1 | 82 A ± 2 |
II | CD | 38.25 | | | 326 a ± 4 | | |
LYS | 35.29 | 5.63 | 4.88 | 250 b ± 4 | 58 A ± 1 | 51 B ± 1 |
THR | 35.61 | 3.74 | 3.22 | 302 b ± 5 | 87 B ± 1 | 94 A ± 2 |
TRP | 35.75 | 1.05 | 0.90 | 277 b ± 6 | 310 ± 7 | 308 ± 6 |
ARG | 34.93 | 5.99 | 5.25 | 276 b ± 6 | 54 ± 1 | 53 ± 1 |
ILE | 35.61 | 3.89 | 3.34 | 321 a ± 8 | 84 B ± 1 | 96 A ± 2 |
VAL | 35.57 | 4.59 | 3.95 | 325 a ± 13 | 71 B ± 1 | 82 A ± 3 |
III | CD | 38.24 | | | 348 a ± 6 | | |
LYS | 38.24 | 5.64 | 4.52 | 284 b ± 12 | 62 ± 1 | 63 ± 3 |
THR | 38.24 | 3.77 | 3.02 | 325 a ± 11 | 92 B± 2 | 108 A± 4 |
ILE | 38.24 | 3.90 | 2.73 | 304 b ± 7 | 89 B± 1 | 111 A± 3 |
VAL | 38.24 | 4.65 | 3.26 | 321 b ± 10 | 75 B± 1 | 99 A± 3 |
In the grower period of Experiment I, only the diluted diets, LYS and THR, yielded a significant decline of parameter
b (
Table 7). Diluted diets, ARG and VAL, obviously tended to respond in the same direction, but according to relatively high standard errors, no significant differences were obtained. However, AA efficiency in diluted diets, TRP and ILE, was significantly enhanced. In Experiment II, due to the elevated lysine supply (
Table 2) for each of the diluted diets, a significant impairment of dietary protein quality
(b) was detected, but only the TRP efficiency in the TRP diluted diet was significantly higher (p = 0.038), as compared to diet, CD. Similar effects on model parameter
b were observed in Experiment III, but due to the stronger dilution of diets, ILE and VAL, the response was more pronounced. In line with Experiments I and II, the efficiency of LYS and VAL was not significantly changed (p > 0.05) following dilution of these AAs.
Table 8 summarizes the results of ideal AA ratios as derived from observed individual AA efficiency dependent on experiment and growth period, respectively. Values in parenthesis indicate calculated data related to results without a significantly confirmed limiting position of corresponding AAs, except ARG and VAL in the grower period of Experiment I. In both of these cases, parameter
b was approximately 10 percent lower compared to CD and, for the VAL diluted diet, near to an acceptable statistical significance (p = 0.056).
Table 7.
Model parameters as derived from N balance studies with control (CD) and AA diluted (DD) diets in the grower period (mean ± SEM *).
Table 7.
Model parameters as derived from N balance studies with control (CD) and AA diluted (DD) diets in the grower period (mean ± SEM *).
Exp. | Diet | N-content (g/kg) | c (g/16g N) | b (b·106) | bc−1 |
---|
CD | DD | CD | DD |
---|
I | CD | 33.73 | | | 473 a ± 26 | | |
LYS | 31.20 | 5.47 | 4.73 | 398 b ± 9 | 86 ± 5 | 84 ± 2 |
THR | 31.47 | 3.55 | 3.04 | 424 b ± 7 | 133 ± 7 | 139 ± 2 |
TRP | 31.59 | 0.99 | 0.85 | 484 a ± 21 | 478 B ± 27 | 572 A ± 25 |
ARG | 30.87 | 5.72 | 5.00 | 431 a ± 18 | 83 ± 5 | 86 ± 4 |
ILE | 31.48 | 3.67 | 3.14 | 465 a ± 13 | 129 B ± 7 | 148 A ± 4 |
VAL | 31.42 | 4.46 | 3.83 | 426 a ± 7 | 106 ± 6 | 111 ± 2 |
II | CD | 35.42 | | | 462 a ± 7 | | |
LYS | 32.62 | 5.63 | 4.89 | 392 b ± 10 | 82 ± 1 | 80 ± 2 |
THR | 32.93 | 3.74 | 3.22 | 408 b ± 8 | 123 ± 2 | 127 ± 3 |
TRP | 33.10 | 1.05 | 0.90 | 424 b ± 10 | 441 B ± 7 | 472 A ± 12 |
ARG | 32.32 | 5.99 | 5.26 | 394 b ± 12 | 77 ± 1 | 75 ± 2 |
ILE | 32.93 | 3.89 | 3.35 | 398 b ± 6 | 119 ± 2 | 119 ± 2 |
VAL | 32.89 | 4.59 | 3.96 | 393 b ± 13 | 101 ± 2 | 99 ± 3 |
III | CD | 35.44 | | | 474 a ± 18 | | |
LYS | 35.44 | 5.64 | 4.52 | 373 b ± 15 | 84 ± 3 | 83 ± 3 |
THR | 35.44 | 3.77 | 3.02 | 419 b ± 12 | 126 B± 5 | 139 A± 4 |
ILE | 35.44 | 3.90 | 2.73 | 373 b ± 14 | 122 B± 5 | 137 A± 5 |
VAL | 35.44 | 4.65 | 3.26 | 358 b ± 30 | 102 ± 4 | 110 ± 9 |
Table 8.
Summarized ideal AA ratios as derived from observed individual AA efficiency related to LYS (LYS = 100).
Table 8.
Summarized ideal AA ratios as derived from observed individual AA efficiency related to LYS (LYS = 100).
Experiment | LYS | THR | TRP | ARG | ILE | VAL |
---|
Starter period |
I | 100 | 60 | (16) * | 100 | (56) * | (70) * |
II | 100 | 61 | 19 | 110 | (60) * | (70) * |
III | 100 | (57) * | nd | nd | 55 | 63 |
Mean ** | 100 | 60 | 19 | 105 | 55 | 63 |
Grower period |
I | 100 | 62 | (15) * | 100 | (58) * | 78 |
II | 100 | 65 | 17 | 110 | 69 | 83 |
III | 100 | 60 | nd | nd | 61 | 77 |
Mean ** | 100 | 62 | 17 | 105 | 65 | 79 |
The observed variability within the experiments and experimental periods was relatively low, as compared to literature data [
10,
12,
17,
22,
59,
60]. All derived IAAR were based on diets with equal native protein sources and constant protein quality over the whole experiment, respectively.
From AA requirement studies in the literature it is indicated that most of the variation in IAAR estimates is provided by differences in bird (genotype, gender, age, performance level [
4,
5,
8,
10,
14,
17,
20,
22,
23,
45,
53,
59,
60]), diet (especially feed ingredients, dietary protein concentration, balance and availability of AA [
4,
5,
8,
10,
46]), response parameter (e.g., BW gain, feed conversion rate, protein deposition, body composition [
8,
22,
42,
44,
45,
46,
47,
48,
54,
56,
57,
60]) and the applied mathematical model for AA requirement assessment [
8,
22,
42,
45,
47]. Even if requirement data are expressed in terms of digestible AA, regardless of total
vs. ileal (praecaecal) or true
vs. apparent digestible, the observed considerable variation of IAAR literature data did not decline [
59]. One explanation is that the concluded AA requirement data are only partly directly related to protein deposition. Additionally, assessment of the praecaecal AA digestibility meets only a part of the total utilization process. AA losses during post-absorptive utilization process, in terms of dietary AA efficiency as reported, need more consideration. Furthermore, attention must also be paid due to the nutritive interaction or antagonism between structurally similar AAs, like LYS and ARG [
35], THR and GLY [
36,
37], TRP and large neutral AAs [
38,
39] and, especially, between the BCAAs, LEU, ILE and VAL [
35,
40], because of their impacts on the dietary AA requirement assessment and, consequently, on the derived IAAR.
The observed ideal THR:LYS ratio was slightly below the assumption derived from literature data (
Table 1), but tended to be higher in older chickens (62
vs. 60). These values are in a similar range as reported by Boorman and Burgess [
11], Austic [
13], Mack
et al. [
18] and Coon [
22]. Lower THR to LYS data (56:100) were published by Baker
et al. [
7]. Likewise, Mack
et al. [
18] and Everett
et al. [
41] obtained data suggesting that the ideal ratio of THR:LYS was lower (57 or 59) than previously reported. Leclercq [
42] established THR:LYS ratios between 59 and 62 to 100 based on a non-linear calculation model and BW gain, feed conversion rate and breast meat percentage as reference criteria. However, according to the “broken line” model, the calculated relative ratios were higher (63 to 65), except for feed conversion rate (only 53.5). Considerably higher values (66 to 74) were obtained in other studies [
41,
43,
44,
45,
46,
47,
48]. The average TRP:LYS ratio of 18:100, as preliminarily recommended, was slightly higher than indicated from the mean of reference data (
Table 1), but it was within the analyzed data pool, varying between 14 and 20 for the dietary needs of TRP relative to LYS (100), and agreed with the current results of Corzo [
49].
Derived ratios of ARG (100 or 110) to LYS (100) were variable, but could be ranged rather in the middle of earlier estimates, indicating a very high variability between 90% and 118% related to LYS, as reviewed by Balnave and Barke [
50]. Due to antagonism between ARG and LYS [
51,
52], the results led to the assumption that the increased ARG:LYS ratio in Experiment II could be related to the elevated dietary LYS content. Furthermore, for both ILE:LYS and VAL:LYS ratios, the current studies did not allow a final conclusion. Compared with the means of the analyzed literature data (
Table 1), both calculated IAAR seem to be lower, especially in the starter period. Actual results [
53] confirm this assumption. Generally, it must be concluded that the published ILE and VAL requirement estimates and derived IAAR are very variable, due to the antagonism of BCAA. If the dietary concentration of one of the BCAA was elevated, a significant increase of the requirements for the other two BCAA was observed [
35,
40,
54]. In contrast, D´Mello [
55] reported that a relatively low dietary ILE level of 0.52% permits satisfactory performance data with dietary LEU and VAL concentrations at 0.98% and 0.63%, respectively. In the majority of studies with growing chicken, especially, the LEU content in diets was very high. Consequently, both the requirements of ILE and VAL and the calculated IAAR tended to be higher [
42,
56,
57,
58].
The currently observed higher relative importance of THR, ILE and VAL in grower compared to starter diets is in line with several results derived from N balance and growth studies on broiler chickens [
8,
13,
17,
20,
23,
53,
59].