Beta-carotene (3a160(a))

Recommended Method

as of:

- Commission Implementing Regulation (EU) 2015/1103

For the determination of Beta- carotene in the feed additive:

- spectrophotometric method based on the - European Pharmacopoeia (Ph. Eur. monograph 1069)

For the determination of Beta-carotene in premixtures and feedingstuffs:

- Reverse Phase High Performance Liquid Chromatography (RP-HPLC) coupled to UV detector

Executive Summary

In the current application authorisation is sought for Beta-Carotene under the category nutritional additives, functional group '3a' vitamins, pro-vitamins and chemically well-defined substances having similar effect, according to the classification system of Annex I of Regulation (EC) No 1831/2003. Specifically, authorisation is sought for the use of Beta-Carotene for all animal species and categories. The active substance and the feed additive is Beta-Carotene, which is a crystalline powder with a purity of at least 96 %. It is produced by chemical synthesis and by fermentation from a strain of Blakeslea trispora. It is intended to be used in premixtures, feedingstuffs and water as a formulated product. According to the applicant typical formulations contain a minimum of 10 % Beta-Carotene. The applicant does not propose any minimum or maximum concentration of the feed additive in feedingstuffs or water.

For the determination of the purity of Beta-Carotene (i.e. the percentage mass fraction of Beta-Carotene in the feed additive), the applicant proposed the European Pharmacopoeia method (Ph.Eur.3^rd, monograph 1069). The CRL-FA considers this method suitable to be used within the frame of official control.

For the determination of Beta-Carotene in premixtures and feedingstuffs the applicant proposed a High Performance Liquid Chromatography (HPLC) method, tested for premixtures and feedingstuffs with concentration ranging from 100 to 2000 mg/kg for premixtures and from 10 to 100 mg/kg in feedingstuffs. The following performance characteristics derived from a three-laboratories intercomparison study were reported by the applicant for premixtures and feedingstuffs:

- a recovery rate of circa 100 %,

- a repeatability relative standard deviation (RSD_r) ranging from 2.9 to 8.9 %,

- a reproducibility relative standard deviation (RSD_R) ranging from 3 to 11.5 %, and

- a limit of detection (LOD) of 0.05 mg/kg for feedingstuffs.

Based on these acceptable performance characteristics the CRL considers this method suitable for the determination of Beta-Carotene in premixtures and feedingstuffs within the concentration range covered by the collaborative study. Therefore the CRL recommends for official control the HPLC method submitted by the applicant, for the determination of Beta-Carotene in premixtures and feedingstuffs.

For the determination of Beta-Carotene in water the applicant did not submit any analytical method nor experimental data, therefore the CRL cannot evaluate nor recommend any method for the determination of the active substance in this matrix.

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.