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The results of 15 experiments realized from 1990 have been used to define more precisely the requirement in Methionine and Sulfur Amino Acids. Results show that the daily requirement for Methionine is related to the bird productivity. If the requirement is expressed in mg per gram of egg mass produced, the requirement appeared independent of the daily egg mass produced. From this analysis, we conclude that the Methionine requirement is not related to the Cystine level. The Methionine requirement is estimated to 7.56 mg in total or 7.11mg in digestible per gram of egg mass produced. The requirement in Sulfur Amino Acids depends mainly of the Cystine level. Excess of Cystine increases the requirement in SAA. Without knowledge of a minimum requirement for the Cystine, we suggest to retain a requirement fixed at 12.10mg in total Sulfur Amino Acids or 10,90mg in digestible Sulfur Amino Acids per gram of egg mass produced.


1. Introduction

Diet formulation and the reduction in dietary protein levels, rendered essential for the achievement of economic goals, and to protect the environment, necessitate a better understanding of amino acid requirements. The concept of ideal protein for layers is limited by our understanding of the «requirements».

The determination of a daily Sulfur amino acid and Methionine requirements has been made by many researchers. The results obtained were dependant on the productivity of the birds.

Just as for the other species, genetic progress has a considerable influence on dietary amino acid concentrations. Over the last 30 years, production to a constant age has increased by more than 30 %, while feed consumption has been reduced by about 10 %. An important consequence of this genetic progress has been a change in the daily amino acid requirements. It has also called into question the practice of phase feeding, since productivity remains high over longer and longer periods. The best birds nowadays have daily egg outputs of over 60g/bird right up to 70 weeks of age and over 58 g/bird for the best flocks at 60 weeks.
As a consequence, the recommendations for daily requirements (table 4) are very variable since they do not take into account the evolution of the bird's productivity.


Table 1: Different Sulfur Amino Acids Recommendations




A review of all the research carried out on amino acid requirements enables us, to specify more precisely the requirements in mg per gram of egg produced and to define the requirement in Sulfur Amino Acids. This method permits us to keep abreast of genetic change while limiting protein wastage and allows the comparison of results from different authors irrespective of the level of production obtained.


2. Materials and Methods

15 experiments realised from 1990 to 2005 with a daily production more than 50 g of egg mass have been used to define the requirement in Sulfur Amino Acids. The conclusion of each researcher has been used to define a requirement in mg per gram of egg mass produced. Each value obtained has been expressed as mg of amino acid per gram of egg produced.

Most of experiments are based on values in amino acids analysed for the total value. To express the results in digestible values, we have recalculated the Methionine and Cystine values according to the European Amino Acid Table (WPSA, 1992) for the Sulfur Amino Acids content, and the INRA raw materials table (2002) for the digestible values, to obtain the digestible amino acid values. In the case of Zollitsch's experiment, the total amino acid values have been recalculated.

The results and the requirement in Amino Acids obtained by each author are mentioned in annex (table 2, 3 and 4).

On average, the comparison between the Sulfur Amino Acids content calculated with European Amino Acid Table show few differences with the values used or analysed by each author. For Methionine and for Methionine+Cystine the difference is less than 0.2% on average. Average Digestible values recalculated from the INRA's coefficient (Raw Materials Table 2002) are the following:

Methionine:  0.9403%
Cystine:  0.840%
Methionine+Cystine:  0.9004%

The effect of a Methionine deficiency on the production cost has been studied in considering that the feed during the laying period represents 60% of the cost of production. We consider also, that the egg mass produced reduces all the other expenses. The impact could be estimated at 40% of the cost of production. This, without including the Methionine supplementation cost.

We have established the cost of production in each experiment for each Methionine level according to the following formula:

Production Cost of the diet (in %) = 60*EMopt/EMlevel + 40*FCRlevel/FCRopt

With
EMopt:  daily egg mass obtained with the optimal diet (result expressed by the author)
EMdiet:  daily egg mass obtained with the different level of Methionine studied
FCRopt:  FCR obtained with the optimal diet
FCRdiet:  FCR obtained with the different level of Methionine studied.


3. Results obtained by each researcher

The conclusion of each researcher (table 5) is synthesised in the following table giving the conclusion of their experiment.


Table 5: Conclusion of the researchers




Most researchers expressed a requirement in Sulfur Amino Acids and claimed for a requirement in Sulfur Amino Acids while other researchers looked at the Methionine level.

The main objective in studying the Sulfur Amino Acids requirement is to answer to these questions. Is Cystine a limiting factor in a standard feed? Which quantity of Methionine is used to satisfy a Cystine requirement? The objectives of this study are to answer to these questions and to define more precisely the Sulfur Amino Acids requirement.


Cystine Requirement


A semi-essential amino acid

Cystine is a semi-essential amino acid, synthesized from Methionine. This can be illustrated (table 6) by an experiment conducted Bertram et al (1992). L-cystine was added to a deficient diet without any effect, demonstrating that Cystine cannot replace Methionine.


Table 6: Effect of DL-Methionine and/or L-Cystine supplementation to a low protein Diet on performance of laying hens, 25-37 weeks of age.




An experiment, by Harms and Russel (1996), shows that birds respond at an increase in the Cystine level and also to a higher level in Methionine (table7). This experiment shows clearly a requirement for Cystine and also for Methionine. However the egg mass produced by the birds in this experiment was quite low.


Table 7: Egg mass produced (g/hen/day) when hens are fed diets with varying levels of Methionine and Cystine. (Exp. 5 from R.H. Harms and G.B. Russel, 1996)




In another experiment, Cao et al (1995), obtained the same poor result with 0.148% and 0.416% of digestible Cystine at same digestible Methionine level 0.143%. The result of their experiment is given in the table 8. Hens need a high requirement in Methionine with a low Cystine level to avoid a body weight loss. Methionine is converted in Cysteine on a molecular basis; consequently 1g of Methionine is converted in 0,8g of Cystine on a weight basis.

Zollitsch et al (1996), tried to define the requirement in Cystine. Their estimation of the digestible requirement is 244 mg of Cystine for a production of 54g. This would give a requirement of 4,52 mg of digestible Cystine per g of egg mass produced (or around 5.3 mg expressed in total in total).


Table 8: Effects of dietary digestible Methionine and digestible Cystine levels on laying performance.





Effect of the Cystine level on the Methionine and Sulfur Amino Acids Requirement

Requirements in Methionine and Methionine plus Cystine obtained by each author are given in the table 3 in the annex. We have expressed these results in mg per gram of egg mass produced (table 3) to establish a relation between the total Cystine level and the total Methionine level and between the total Cystine level and the total Methionine+Cystine level (table 4).

The correlation (figure 1) obtained for the requirement between the Cystine level and Methionine and Methionine+Cystine are:

Total M+C requirement (mg/g)   =  +1.09* Cys. + 7.07 (R² = 0.83)
Total Meth. requirement (mg/g) =  -0.096* Cys. + 7.08 (R² = 0.04)

With Cys. expressed in mg of total Cystine per gram of egg mass produced.

These results show clearly the absence of relation between the requirement in Methionine and the Cystine level. This is due to the fact that in most experiments the level of Cystine was sufficient to cover the requirement in Cystine. High feed Cystine values were found in the experiments of Balnave et al (2000), Bertram et al (1995) and Cao et al (1992) without effect in the Methionine requirement expressed in mg per gram of egg. Conversely, the Sulfur Amino Acids requirement appear correlated to the feed Cystine level used in the different experiments.


Figure 1: Effect of the total Cystine Level on the total Methionine and Sulfur Amino acids requirement expressed in mg per g of egg mass produced by each author in 15 experiments made from 1990 to with a daily egg mass produced above 50g.




A feed high in Cystine can contribute to a Methionine deficiency if the constraints in formulation are based only on Methionine+Cystine. The absence of a relationship between the Methionine requirement and the level of Cystine forces the nutritionist to impose a minimum constraint on Methionine. A feed low in Cystine increases the requirement in Methionine and obliges the nutritionist to use a minimum constraint on Sulfur Amino Acids.


4. Methionine Requirement


Results

The average of the 15 experiments in the annex (table 4) gives a requirement expressed in mg per gram of egg mass for:

Total Methionine:  7.56mg ± 0.37
Dig. Methionine:  7.11mg (mean of the digestibility 0.940)

The absence of correlation between the Cystine level and the Methionine leads to the conclusion that the nutritionist has to formulate feed on this constraint (figure 1). Expression of the requirement in mg of gram of egg mass and the study of a lot of experiments allow us to close the debate on the interest to formulate the feed in using Methionine as a constraint.


Figure 2: Influence of the egg mass produced on the daily Methionine requirement and on the requirement expressed in mg to produce 100g of Egg Mass.




The value of 7.56 mg/g appears independent of the egg mass produced. Figure 2 shows the relation between the bird’s productivity and the requirement to produce 1g of egg mass and also, with the daily requirement. The coefficients of regression obtained are 0.55 between the daily egg mass produced and the total Methionine requirement and 0.01 between the daily egg mass produced and the total Methionine requirement expressed in mg to produce 1g of egg mass.

This result establishes clearly the necessity to express a requirement in Amino Acids in mg per gram of egg mass produced.

The use of the results of 15 experiments allows us to define with a good precision the requirement in Methionine. The coefficient of variation obtained for the requirement is only 4.5%.


Cost of Production

The cost of production has been calculated according the formula given in the ‘materials and methods’ chapter. Regression between the level of deficiency and the cost of production is high (over 0.8). Figure 2 shows the relation for a deficiency less 30%.

The relation appears linear and a Methionine reduction of 1% increases the egg cost of production by 0.24%. For a level of Methionine comprised between 70 and 100%, we found this relation:

Production Cost = -0.247x + 124.7 (r²=0.88)
with x= level of deficiency

The cost of the Methionine added is not included, but could be considered low. A 10% variation of the Methionine level affects the cost of production by around 0.5 %, consequently the formula becomes:

Cost = -0,20x + 120 (r²=0.88)

This means that a deficiency of 10% increases the cost of production by 2%.


Figure 3: Relation between the cost of production and the Methionine level of deficiency





Figure 4: Relation between the cost of production and the Methionine level of deficiency with a deficiency less than 30%. 




Influence of a Methionine deficiency on performances

A Methionine deficiency affects all production parameters, egg weight, egg number and feed efficiency. It is interesting to see the relation between the feed consumption and the Methionine deficiency (figure 5). Birds increase generally their consumption in case of Methionine deficiency; consequently a deficiency has a strong effect on the feed consumption ratio (figure 6). This effect has been observed in a previous study, (Joly, 1995) and has not been observed for other amino acids.

Egg weight and percentage of lay seem affected in the same proportion.


Figure 5: Effect of the level of Methionine in percentage of the author’s requirement on the Egg Mass produced and the daily feed consumption.





Figure 6: Effect of the level of Methionine in percentage from 55 to 100% of the author’s requirement on the Feed Conversion Ratio expressed in percentage.




It is well known that a Methionine deficiency is also responsible for feather pecking and cannibalism. This is illustrated in the Dànner’s experiment:


Table 9: Effects of Sulfur Amino Acids level on feather pecking and liveability.




5. Methionine and Sulfur amino acids Requirement


Is there a requirement in Sulfur Amino Acids?

The main objective for the feed formulator is to produce a “balanced” feed at the least cost. Minimum of constraints is always costly. It is important to know if a minimum of constraint in Sulfur Amino Acids is need. We have found that the requirement in Methionine doesn’t depend of the Cystine level (figure 1). However the coefficient of regression between the Cystine level in the feed and the requirement in Sulfur Amino Acids is very high (r²=0.83). This means that the requirement for Sulfur Amino Acids depends of the feed Cystine level. It appears, there is only a Methionine requirement.

To precise this point we compared the results of the experiments in which the Cystine level was under or above 5 mg per gram of egg produced (table5).

The selection of the experiments according the Cystine level (mg/g) doesn’t allow us to see any difference for the requirement in Methionine. It appears that the requirement for Cystine is lower than the value estimated by Zollitsch et al (1996).


Table 5: Influence of the Cystine level on the Methionine and Sulfur Amino Acids requirement




Which Sulfur Amino Acids level to use?

To determine a requirement in Cystine, we made a regression between the Cystine level and the Methionine level (figure 7). From this result, it would be possible to conclude that the requirement in Cystine is fewer than 4.5 mg per gram of egg produced.

Harms and Russel (1996) made the following conclusion: “A corn-soybean meal diet would always furnish more Cystine than required by the commercial laying hen; therefore, it is not necessary to formulate for SAA requirement”.

However, Zollitsch et al (1996) have estimated the digestible Cystine requirement at 4.5 mg/g (approximately 5.3 mg/g expressed in total Cystine). The risk of a Cystine deficiency exits with the use of some specific raw materials having a low Cys./Met. ratio, like Fish Meal, Meat and Bone Meal or Sunflower Meal. For this reason we think it is important to fix a minimum of Sulfur Amino Acids. Looking at the results of experiments in which the level of Cystine was low, we think reasonable to use a safety margin and to fix the minimum in total Sulfur Amino Acids at 12.10 mg per gram of egg produced. This value is the average of the 7 experiments having the lowest requirement in Sulfur Amino Acids.


Figure 7: Effect of low total Cystine Level (above 5mg/g) on the total Methionine and Sulfur Amino acids requirement expressed in mg per g of egg mass.




6. Conclusions


Requirement per gram of egg produced

The estimated requirements given by each author of the 15 experiments, expressed in mg of amino acids per gram of egg mass are:

Methionine (total):  7.56 mg/g
Methionine + Cystine (total): 12. 1 mg/g

The estimation of the digestible value has been made in using the digestible coefficient expressed in the raw material table published by INRA (2002). The digestible value (Methionine added included) from 12 experiments has been calculated and are 94.0 % for Methionine, 84.0 % for Cystine and 90.0 % for Sulfur Amino Acid. Consequently the estimation of the requirement in digestible values are:

Dig. Methionine:  7.10mg/g
Dig. Methionine + Cystine: 10.90 mg/g


Requirement per Day

With a daily production at the peak of production fixed at 59.5g per day, the daily requirement could be estimated at:

Methionine (total):  450 mg
Methionine + Cystine (total):  720 mg
Dig. Methionine:  420 mg
Dig. Methionine + Cystine:  650 mg

These results are similar at those found in our previous review, Joly (1999), for Methionine but lower for the Methionine plus Cystine requirement.


Requirement according to the Feed Efficiency

Another way is to express the requirement in percentage of amino acids per kg of feed, according to the feed conversion ratio. The following table gives the percentage of Sulfur Amino Acids according to the feed conversion during the peak production period.





Requirement at start of lay

Due to a lower body weight and consequently a lower feed consumption, from 18 to 28 weeks, we advice to increase the amino acids by 6 % during this period to satisfy the birds requirement (Joly, 2002).


References

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Table 2. Effect of Methionine and S. Amino acids level on performances and estimation of the production cost established from 15 experiments made from 1990 with a daily egg mass produced above 50g.


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Table 3. Effect of Methionine and Sulfur Amino acids level on performances and estimation of the requirement established in 15 experiments made from 1990 with a daily egg mass produced above 50g.


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Table 4. Methionine and Sulfur Amino acids requirement expressed in mg per g of egg mass produced by each author, and estimated from the European Amino Acid Table (WPSA, 1992) and the INRA raw materials table (2002) for the digestible values in 15 experiments made from 1990 with a daily egg mass produced above 50g.


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