6-phytase EC 3.1.3.26 (4a23)

Recommended Method

as of:

- Commission Implementing Regulation (EU) 2016/329

For the quantification of 6-phytase activity in the feed additive:

- colorimetric method based on the enzymatic reaction of phytase on the phytate - VDLUFA Method Book, Vol. III, 27.1.1

For the quantification of 6-phytase activity in premixtures and mineral feeds:

- colorimetric method based on the enzymatic reaction of phytase on the phytate - VDLUFA Method Book, Vol. III, 27.1.3

For the quantification of 6-phytase activity in feedingstuffs:

- colorimetric method based on the enzymatic reaction of phytase on the phytate - EN ISO 30024

Executive Summary

In the current application authorisation is sought under article 4(1) for ENZY PHOSTAR®, under the category/functional 4(a) "zootechnical additives"/"digestibility enhancers" according to the classification system of Annex I of Regulation (EC) No 1831/2003.

Specifically, authorisation is sought for the use of the feed additive for chickens and turkeys for fattening, chickens reared for laying, turkeys reared for breeding, laying hens, minor avian species including laying birds (game birds, geese, pigeons, sporting and ornamental birds), weaned pigs, pigs for fattening, sows for reproduction and minor porcine species. According to the Applicant, the active agent of ENZY PHOSTAR® is 6-phytase (EC 3.1.3.26), produced by fermentation of a genetically modified strain of the yeast Pichia pastoris (DSM 25375). The Applicant uses the enzyme activity unit as defined in the EN ISO 30024, where one phytase unit (U) is the amount of enzyme which releases one micromole of inorganic phosphate from sodium phytate per minute at pH 5.5 and 37 ^oC. The product is intended to be marketed in a powder and liquid formulations having a guaranteed minimum phytase activity of 40000 U/g. The carrier in the solid formulation is starch, while glycerol, sodium chloride and sodium benzoate, are used for the liquid formulation. The feed additive is intended to be included through premixtures to obtain a minimum activity in feedingstuffs of 125 U/kg (for laying hens and minor avian species for laying) and 250 U/kg (for the other species of interest).

For the determination phytase activity in the feed additive, the Applicant submitted the ring trial validated colorimetric method (VDLUFA 27.1.1) measuring the inorganic phosphate released by the enzyme from the sodium phytate substrate. Based on the acceptable performance characteristics, the EURL recommends for official control the VDLUFA 27.1.1 method for the determination of phytase activity in the feed additive.

The Applicant proposed the ring-trial validated colorimetric method (VDLUFA 27.1.2) for the determination phytase activity in premixtures and feedingstuffs. Upon request from the EURL, the Applicant provided additional experimental data demonstrating the applicability of the internationally recognised ring-trial validated colorimetric EN ISO 30024 method. Based on the experimental evidence available the EURL recommends for official control the EN ISO 30024 method for the determination of phytase activity in feedingstuffs.

Furthermore, the EURL identified and recommends the newly available ring-trial validated colorimetric method (VDLUFA 27.1.3) for the determination of phytase activity in mineral feeds and premixtures, aiming to extend the scope of the EN ISO 30024 to these matrices.

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) is not considered necessary.