Riboflavin from Eremothecium ashbyi as a dried fermentation product (≥ 5 %): HPLC-FLD, VDLUFA and EN 14152 — EFSA recorded dioxins above the feed material limit in two batches and fumonisin B1 up to 2000 µg/kg
Commission Implementing Regulation (EU) 2025/1254, published on 26 June 2025, authorises until 16 July 2035 riboflavin (vitamin B2, 3a827) produced with the non-genetically modified strain Eremothecium ashbyi CCTCCM 2019833 in the form of a dried, inactivated fermentation product with at least 5 % riboflavin and moisture ≤ 7 % for all animal species. In 2021 EFSA could not conclude on safety; in 2024 — after new genotoxicity studies and a 90-day study — it found the additive safe.
- Methods in the annex: riboflavin in the additive — HPLC-FLD or HPLC-UV to VDLUFA Bd. III, 13.9.1; in premixtures — HPLC-UV (VDLUFA); in compound feed as total vitamin B2 — HPLC-FLD to EN 14152. EURL report (2021 opinion): HPLC-FLD for the additive — RSDr 2.4 %, RSDip 2.7 %, recovery 98 %; VDLUFA in premixtures (868–15 990 mg/kg) — RSDr 2.4–4.7 %, RSDR 4.2–7.3 %, recovery 86–100 %; EN 14152 (acid hydrolysis) — RSDR 7.3–7.9 %.
- Five batches (2021 opinion): 6.5–7.4 % riboflavin (mean 6.9 %), crude protein 38–41 %, 20–26 % dry microbial biomass. PCDD/F dioxins: ≤ 0.73 ng WHO-TEQ/kg in three batches but 1.63 and 1.77 ng in two — above the maximum content of Regulation (EU) No 277/2012 for feed materials; fumonisin B1 852–2000 µg/kg; B. cereus 1400–86 000 CFU/g in pilot batches and at most 200 CFU/g in nine later ones. EFSA recommended attention to these parameters in production; the act sets no limits for them.
- Opinion of 15 October 2024: in vitro micronucleus test (OECD TG 487) of limited relevance; combined comet assay (OECD TG 489) and micronucleus test (OECD TG 474) in rats up to 6000 mg/kg bw per day — no genotoxicity (an increase in %TI only in the ileum at the mid dose was judged not biologically relevant). 90-day feeding study: NOAEL 240 000 mg additive/kg feed, i.e. 21 600 mg/kg bw per day — the highest dose tested.
- From the NOAEL (uncertainty factor 100) EFSA calculated safe additive concentrations of 2406–42 240 mg/kg complete feed — above the recommended doses of 60–1700 mg/kg (e.g. 160–200 mg/kg for starter chickens, 800–1600 mg/kg for shrimp). The additive is not a skin or eye irritant nor a skin sensitiser, but is a respiratory sensitiser — the act requires respiratory protection where needed.