Preparation of 6-phytase (EC 3.1.3.26) - 4a37

In the current application, authorisation of a preparation of 6-phytase (EC 3.1.3.26) is sought under Article 4 for all poultry and all pig species under the category/functional group 4(a) "zootechnical additives"/"digestibility enhancers" according to Annex I of Regulation (EC) No 1831/2003. 

According to the Applicant, the active agent of the product is 6-phytase, produced by fermentation of the genetically modified strain Trichoderma reesei CBS 146250. Other preparation of 6-phytase from a different Trichoderma reesei strain is currently authorised as feed additive. 

The activity of 6-phytase is expressed in phytase units (FTU) where "one FTU is the amount of enzyme which releases one micromole of inorganic orthophosphate from a sodium phytate substrate per minute at pH 5.5 and 37 °C". This definition is in agreement with the phytase activity unit as described in EN ISO 30024. 

The product is intended to be marketed in three different formulations namely Axtra® PHY GOLD 30 L (liquid), Axtra® PHY GOLD 30 T (granular and thermostable) and Axtra® PHY GOLD 65 G (granular) with a guaranteed minimum 6-phytase activity of 30000 FTU / g for the Axtra® PHY GOLD 30 L and the Axtra® PHY GOLD 30 T formulations and 65000 FTU / g for the Axtra® PHY GOLD 65 G formulation. The product is intended to be included through premixtures or directly in feedingstuffs to obtain a minimum recommended activity of 300 FTU / kg feedingstuffs for laying hens and other birds for laying and 500 FTU / kg feedingstuffs for the other target species. 

The Applicant submitted single-laboratory validated and further verified methods for the quantification of the phytase activity in the product (Axtra® PHY GOLD), premixtures and feedingstuffs. The submitted methods are very similar to the ring-trial validated EN ISO 30024 method. 

Upon request of the EURL, the Applicant compared both protocols confirming that equivalent results are obtained when applying the slightly different methods to feedingstuffs containing the product (Axtra® PHY GOLD). 

Additionally the EURL is aware of other ring-trial validated VDLUFA methods specifically describing the preparation of premixtures (VDLUFA 27.1.3) and feed additives (VDLUFA 27.1.4) for the quantification of their phytase activity according to EN ISO 30024. 

Based on the performance characteristics available the EURL recommends for official control the ring-trial validated EN ISO and VDLUFA colorimetric methods mentioned above for the quantification of the phytase activity in the product, premixtures and feedingstuffs.

Further testing or validation of the methods to be performed through the consortium of National Reference Laboratories as specified by Article 10 (Commission Regulation (EC) No 378/2005, as last amended by Regulation (EU) 2015/1761) is not considered necessary.

Recommended Method

as of:

-Commission Implementing Regulation (EU) 2023/1167

For the quantification of phytase activity in the feed additive: colorimetric method based on the enzymatic reaction of phytase on phytate – VDLUFA 27.1.4

For the quantification of phytase activity in premixtures: colorimetric method based on the enzymatic reaction of phytase on phytate – VDLUFA 27.1.3

For the quantification of phytase activity in compound feeds: colorimetric method based on the enzymatic reaction of phytase on phytate – EN ISO 30024