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The substance Beta-Cyclodextrin has been tested on the 5 strains of Salmonella typhimurium (TA 98, TA 100, TA 1535, TA 1537, TA 1538) with and without metabolic activation (S9 -mix) according to the protocol of Ames and similar to OECD Guideline 471. A range of non-toxical concentrations had been determined in a preliminary experimentation on the strain TA 100 without metabolic activation.The 5 concentrations chosen (0.1, 0.5, 1, 2 and 4 mg/plate) have been tested in triplicate on the 5 strains quoted above, with and without metabolic activation. The results were confirmed in a second experiment separately from the first one.In each test a negative control (solvent) and a positive one (specific standard mutagene) had been included. The values obtained were consistent with the standards. Under these circumstances, the product did not show any mutagene power towards thestrains TA98, TA 100, TA 1535, TA 1537, TA .1538 with and withoutmetabolic activation.

A study equivalent to OECD 476 was performed to investigate the potential of beta-Cyclodextrin (100% pure) to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. The assay was performed in two independent experiments, using two parallel cultures each. Both main experiments were performed with and without liver microsomal activation (S9) and a treatment period of 3 hours.The highest concentration of the test item was 1000 µg/mL. The tested concentrations were 30 -100 -300 and 1000µg/mL for both experiments with and without metabolic activation. No substantial and reproducible dose dependent increase of the mutation frequency was observed in both main experiments. Appropriate reference mutagens, used as positive controls, induced a distinct increase in mutant colonies and thus, showed the sensitivity of the test item and the activity of the metabolic activation system. In conclusion it can be stated that under the experimental conditions reported the test item did not induce gene mutations at the HPRT locus in V79 cells. Therefore, beta-Cyclodextrin is considered to be non-mutagenic in the HPRT assay.

Beta-Cyclodextrin was also assessed in an in vitro Chromosome Aberration assay with human lymphocytes similar to the OECD guideline 473. The substance is soluble in aqueous medium, solutions were prepared in RPMI 1640 at the maximum concentration generally used at the laboratory, ie., 10 mg/ml. This solution was added at 10 % in culture medium. The highest final

concentration was 1000 µg/ml. A Preliminary test on cytotoxicity showed a low cytotoxicity of beta-cyclodextrin both with and without metabolic activation, at the highest studied dose 1000 µg/ml, demonstrated by a weak mitotic index decrease. The tests were performed with and without metabolic activation (S9 -mix) at concentrations of 100, 300 and 1000 µg/ml with 1 (with metabolic activation) and 24 hr (without metabolic activation) exposition time. Solvent, negative and positive controls worked as expected. Reference clastogens (mitomycin C without metabolic activation and cyclophosphamide with metabolic activation) were tested in order to demonstrate the sensitivity of the cells and the effectiveness of the metabolic activation system. Both tests with and without metabolic showed no dose dependent positive results presents and thus the test substance did not show clastogenlc

activity in the in vitro human lymphocyte metaphase analysis test.

The recessive lethal mutation inducing effect of the compound beta-cyclodextrin was studied in Drosophila melanogaster by sex-linked recessive lethal test SLRL. Newly hatched Oregon-R males were treated with 1,6; 8.0 and 16 mM of beta-cyclodextrin for 24 hours. Each F1 female represents one paternal X-chromosome, treated in the male gametes. The vials to breed the F2 generation each corresponds to one treated male gamete and for that reason never should contain more than one F1 female. The vials of the F2 generation are then individually inspected for the presence of wild type /round red eyes/ males. Spontaneous (control) mutation frequency was determined with untreated animals. Suitability of test system was controlled by the positive effect of methyl methanesulfonate (MMS) a well-known mutagenic agent. It can be stated that the compound under study did not increase the frequency of the sex-linked recessive lethal mutation above the spontaneous level in the Drosophila melanogaster.

The mutagenic potential of the test article beta-Cyclodextrin (purity not specified) was tested in the I.O.P.S., OFl (IFFA CREDO) mouse using the micronucleus test, at a preliminarily defined dose of 100mg/kg according to a procedure similar to OECD 474.

The animals (5 males and 5 females per group) were administered the product by the intraperitoneal route (vehicle 1% carboxymethylcellulose hydrogel) and killed 24, 48 or 72 hours after administration. For each animal an examination of 1000 polychromatic erythrocytes obtained from the femoral bone marrow was performed.The statistical analysis of the results does not show any significant increase in the number of polychromatic erythrocytes bearing micronuclei in the animals treated with the test substance. The results obtained with the positive control (cyclophosphamide) are significantly positive.Concerning the ratio normochromatic/polychromatic erythrocytes which in the absence of toxical effect is close to 1, there was no significant increase in this value in the animals treated with the test article. A significant increase was observed in the animals treated with the positive control.To conclude, the test article did not induce any mutagenic effect in the mouse when administered at a dose of 100 mg/kg



Short description of key information:
All in vivo (micronucleus test) and in vitro (chromosome aberration, HPRT, Ames) Tests performed are negative.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

The substance did not show any mutagenic or clastogenic potential in any of the tests performed.