Safety evaluation of a mixture of the human-identical milk oligosaccharides 2′-fucosyllactose and difucosyllactose

Baldwin

et al. 2019

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Human breastmilk is a mixture of nutrients, hormones and bioactive molecules that are vital for infant growth and development. Infant formula (IF) lacks many of these compounds, most notably human milk oligosaccharides (HMOs), which are abundant in breastmilk but scarce in IF. Sialyllactoses, such as 3′‐sialyllactose, constitute a large portion of the HMO fraction. To produce IF that matches breastmilk more closely, biosynthesized human‐identical milk oligosaccharides (structurally identical to HMOs) such as 3′‐sialyllactose sodium salt (3′‐SL) are proposed for use in IF and foods for the general population. The safety assessment of 3′‐SL comprised in vitro genotoxicity tests and a 90‐day oral (gavage) toxicity study. This is the first 90‐day study conducted with 3′‐SL using neonatal rats (7 days old at the start of dosing—equivalent age to newborn human infants in terms of central nervous system and reproductive development), demonstrating the safety of 3′‐SL for consumption by infants, the most sensitive age group. The neonatal rats received 3′‐SL at doses up to 5,000 mg/kg body weight (BW)/day and reference controls received 5,000 mg/kg BW/day of fructooligosaccharide (an ingredient approved for use in IF) for comparison with the high‐dose 3′‐SL group, followed by a 4‐week recovery period. There was no evidence of genotoxicity in vitro. In the absence of any test item‐related adverse effects in the 90‐day study, the high dose (5,000 mg/kg BW/day) was established as the no‐observed‐adverse‐effect level. This confirms the safety of 3′‐SL for use in IF for infants, as well as in functional foods for the general population.

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Toxicological safety assessment of the human‐identical milk oligosaccharide 3′‐sialyllactose sodium salt

Baldwin

et al. 2019

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“…The method for the preclinical safety assessment of 6′‐SL (two in vitro genotoxicity tests and a 90‐day oral toxicity study) is similar to that used for other HiMOs, such as 3′‐sialyllactose sodium salt (3′‐SL) (Phipps et al, , accepted manuscript), LNnT (Coulet et al, ), 2′‐FL (Coulet et al, ), lacto‐ N ‐tetraose (Phipps, Baldwin, Lynch, Stannard, et al, ) and 2′‐FL/DFL (Phipps, Baldwin, Lynch, Flaxmer, et al, ). The in vitro assays represent the standard genotoxicity test battery covering gene mutations and chromosome aberrations using the minimum recommended number of tests (EFSA, ) and the 90‐day oral toxicity study was conducted to assess the potential subchronic toxicity of not only 6′‐SL itself, but of ‘other carbohydrates' within the final product (see Table ).…”

Section: Discussionmentioning

confidence: 99%

“…Duplicate cultures were prepared for each concentration and positive control, and quadruplicate cultures were prepared for vehicle controls. Subsequent treatment/handling of cells, statistical analyses and evaluation of the assay results were performed as previously described by Phipps, Baldwin, Lynch, Flaxmer, et al ().…”

Section: Methodsmentioning

confidence: 99%

“…A previous publication on the safety of enzymatically manufactured 6′‐SL differed in several aspects (Gurung et al, ) from the approach used in this study, which is similar to that used for other human‐identical milk oligosaccharides (HiMOs) such as lacto‐ N ‐neotetraose (LNnT) (Coulet, Phothirath, Constable, Marsden, & Schilter, ), 2′‐fucosyllactose (2′‐FL) (Coulet, Phothirath, Allais, & Schilter, ), a mixture of 2′‐fucosyllactose and difucosyllactose (2′‐FL/DFL) (Phipps, Baldwin, Lynch, Flaxmer, et al, ), and lacto‐ N ‐tetraose (Phipps, Baldwin, Lynch, Stannard, et al, ). This approach included conduct of a 90‐day oral gavage study in neonatal rats [dosed from postnatal day (PND) 7] designed to simulate human infant exposure and allow for assessment of any potential systemic toxicity following exposure covering pre‐ and post‐weaning from growth into adulthood.…”

Section: Introductionmentioning

confidence: 99%

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Toxicological safety evaluation of the human‐identical milk oligosaccharide 6′‐sialyllactose sodium salt

Baldwin

et al. 2019

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Human milk oligosaccharides (HMOs) are abundant in breastmilk, but their presence in infant formula is negligible. Sialylated HMOs, such as 6′‐sialyllactose, constitute a significant portion of the HMO fraction of human milk and are linked to important biological functions. To produce infant formula that is more comparable with human milk, biosynthesized sialyllactoses known as human‐identical milk oligosaccharides (structurally identical counterparts to their respective naturally occurring HMOs in breastmilk) are proposed for use in infant formula and other functional foods for the general population. To support the safety of 6′‐sialyllactose sodium salt (6′‐SL), a 90‐day oral (gavage) toxicity study and in vitro genotoxicity tests were conducted. The 90‐day study is the first to be conducted with 6′‐SL using neonatal rats (day 7 of age at the start of dosing), thus addressing safety of 6′‐SL for consumption by the most sensitive age group (infants). In the 90‐day study, neonatal rats received 6′‐SL at doses up to 5000 mg/kg body weight (BW)/day and reference controls received 5000 mg/kg BW/day of fructooligosaccharide (an ingredient approved for use in infant formula) for comparison with the high‐dose 6′‐SL group, followed by a 4‐week recovery period. There was no evidence of genotoxicity in vitro. No test item‐related adverse effects were observed on any parameter in the 90‐day study, thus the high dose (5000 mg/kg BW/day) was established as the no‐observed‐adverse‐effect level. These results confirm that 6′‐SL is safe for use in formula milk for infants and in other functional foods for the general population.

Genotoxicity and neonatal subchronic toxicity assessment of a novel mixture of the human‐identical milk oligosaccharides lacto‐N‐fucopentaose I and 2′‐fucosyllactose

Stannard

et al. 2020

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Human milk oligosaccharides (HMOs) are a complex group of bioactive molecules largely observed in human breast milk but also occurring in limited amounts in other mammalian milks. Advances in biotechnology have enabled production of humanidentical milk oligosaccharides (HiMOs), structurally identical molecules to HMOs found naturally in human milk, intended for addition to infant formula to more closely replicate breast milk. Biosynthesis of a novel mixture of two major HMOs, lacto-N-fucopentaose I and 2 0-fucosyllactose (LNFP-I/2 0-FL), recently became possible. To support the safety of LNFP-I/2 0-FL for use in infant formula and other foods, it was subject to a safety assessment comprising a bacterial reverse mutation test, an in vitro mammalian cell micronucleus test, and a 90-day oral gavage study in neonatal rats. In the 90-day study (the first HiMO study to include the new endocrinesensitive endpoints described in the 2018 version of OECD Test Guideline 408), LNFP-I/2 0-FL was administered by oral gavage to neonatal rats once daily (from Day 7 of age) for 90 consecutive days, at doses up to 5000 mg/kg bw/day, followed by a 4-week recovery period. Concurrent reference controls received 5000 mg/kg bw/day of the approved infant formula ingredient oligofructose. LNFP-I/2 0-FL was nongenotoxic in vitro. The highest dose tested (5000 mg/kg bw/day) was established as the no-observed-adverse-effect level in the 90-day study, as there were no test article-related adverse effects on clinical observations, body weight, food consumption, clinical pathology, and organ weights nor any noteworthy macroscopic or microscopic findings. This supports the safety of LNFP-I/2 0-FL for its intended uses in food.