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Description of key information

- As the test substance is an enzyme (i.e. proteins consisting of sequences of amino-acids), it is expected to be denaturated and digested in the gastrointestinal tract, just like other food proteins.

- No substantial dermal absorption is expected based on the large molecular weight (>> 500) and the partition coefficient ( logPow< -1) of the test substance

- As the test substance is sensitising to the respiratory tract, it is assumed to be inhaled and potentially absorbed.

Key value for chemical safety assessment

Additional information

Basic toxicokinetics

As there are not direct studies on the basic toxicokinetic properties of the test substance, a justification based on available data of the test substance as well as structural analogues is provided. Available data include physico chemical properties and toxicological profiles. The structural analogues are used in the read across justification for the test substance. Since all source substances and the target substance are enzymes, i.e. proteins, consisting of sequences of amino-acids, it is known that they are denaturated and digested in the gastrointestinal tract, just like other food proteins. Metabolites will be absorbed via the gastrointestinal tract, but there will be no systemic availability as a complete enzyme (Olempska-Beer et al., 2006; Basketter et al., 2012). The physicochemical characteristics, as far as known, indicate that enzymes are not expected to have substantial dermal absorption, since they are very large (Molecular weights >> 500) and have a partition coefficient (log Pow) below -1 (Guidance on information requirements and chemical safety assessment Chapter R.7.c: Endpoint specific guidance, R.7.12 Guidance on Toxicokinetics). Based on the limited information, estimation of the absorption via inhalation is difficult. As several of the substances discussed in this document are considered to be sensitizing to the respiratory tract, these substances are assumed to be inhaled and potentially absorbed. As no acute inhalation toxicity is observed it is impossible to determine the amount that is absorbed.


Toxic mechanism (mode of action)

The main action of the target substance and source substance is to act as a biocatalyst in the hydrolysis 1-->4 or 1-->3 linkages in various substrates, mostly glucans and similar materials. Enzymes, in general, are proteins which catalyse chemical reactions. Every cell in living organisms contains enzymes that transform chemicals. Hydrolytic enzymes, such as the target and source substances, decompose macromolecules into small units (Pariza and Foster, 1983). The difference in function of different enzymes is largely caused by the differences in shapes of the enzymes, which causes certain substrates to bind to an active site of the enzyme. The active sites depend on the tertiary structure of the enzyme. This largely explains why e.g. cellulase catalyses the hydrolysis of 1->4 linkages in beta-D-glucans, but not of 1->3 linkages (which are catalysed by endo-1,3(4)-ß-glucanase). However, some hydrolytic enzymes work on multiple linkages, such as the endo-1,3(4)-ß-glucanase, which hydrolyses (in certain cases), 1->3 and 1->4 linkages. Denaturation and digestion of the enzymes, which is expected to occur in the gastro-intestinal tract, will lead to loss of the enzymatic activity, because the tertiary structure of the enzyme is destroyed.

Chemically, the reaction mechanism of the hydrolytic enzymes, such as endo-1,4-ß-glucanase, cellulase (which is also an endo-1,4-ß-glucanase), endo-1,3(4)-ß-glucanase, xylanase, hemicellulase and α-amylase is very similar. They all hydrolyse O- and S-glycosyl compounds. Because of this similarity in chemical reactions of the various hydrolytic enzymes, it can be expected that any toxic mode of action will also be similar. Pariza and Foster (1983) refer to exhaustive literature reviews by the FDA to conclude that food enzyme preparations from microbial and non-microbial sources are not toxic and not pathogenic and safe to consume. There are also numerous notifications to the FDA on enzymes considered to be ‘generally recognized as safe’ (GRAS). In the studies on toxicity on the target substance and the source substances, the only relevant effects seen are skin sensitisation and respiratory sensitisation. Basketter et al. (2012) have summarised what is known or suspected in relation to why enzymes are allergens. There is not a complete picture why only a limited number of proteins actually act as respiratory allergens. It is considered, that the TH2 subpopulation of helper cells is involved. Perhaps the enzymatic characteristics of enzymes also impact their immunological effects. However, Basketter et al. (2012) finally concludes that “However, at present it is not possible to confirm or refute whether specific types of enzymes action are always directly linked to allergenicity.”