With the recent concern regarding cellulose enhancing radionuclide mobility upon its degradation to ISA, disposal of cellulosic wastes is being held off until the disposal safety is vindicated. Thus, a rational assessment should be conducted, applying an appropriate cellulose degradation model considering the disposal environment and cellulose degradation mechanisms. In this paper cellulose degradation mechanisms and the disposal environment are studied to propose the best-suitable cellulose degradation model for the domestic 1st phase repository. For the cellulose to readily degrade, the pH should be greater than 12.5. As in the case of SKB, 1BLA is excluded from the safety assessment because the pH of 1BLA remains below 12.5. Furthermore, despite cellulose degradation occurring, it does not always produce ISA. At low Ca2+ concentration, the ISA yield rate is around 25%, but at high Ca2+ concentration, the ISA yield rate increases up to 90%. Thus, for the cellulose to be a major concern, both pH and Ca2+ concentration conditions must be satisfied. To satisfy both conditions, the cement hydration must be in 2nd phase, when the porewater pH remains around 12.5 and a significant amount of Ca2+ ion is leaching out from the cement. However, according to the safety evaluation and domestic research, 2nd phase of cement hydration for silo concrete would achieve a pH of around 12.4, dissatisfying cellulose degradation condition like in 1BLA. Thus, cellulose degradation would be unlikely to occur in the domestic 1st phase repository. To derive waste acceptance criteria, a quantitative evaluation should be conducted, conservatively assuming cellulose is degraded. To conduct a safety evaluation, an appropriate degradation model should be applied to determine the degradation rate of cellulose. According to overseas research, despite the mid-chain scission being yet to be seen in the experiments, the degradation model considering mid-chain scission is applied, resulting in an almost 100% degradation rate. The model is selected because the repositories are backfilled with cement, achieving a pH greater than 13, so extensive degradation is reasonably conservative. However, under the domestic disposal condition, where cellulose degradation is unlikely to occur, applying such model would be excessively conservative. Thus, the peeling and stopping model derived by Van Loon and Haas, which suggests 10~25% degradation rate, is reasonably conservative. Based on this model, cellulose would not be a major concern in the domestic 1st phase repository. In the future, this study could be used as fundamental data for planning waste acceptance criteria.

Cellulose-based wastes can be degraded into short-chain organic acids at the cementitious radioactive waste repository. Isosaccharinic acid (ISA), one of the main degradation products, can form the chelate complex with metals and radionuclides, and these complexes have a potential that can accelerate to move the radionuclides to far-field from the repository. This study characterized the amount of generated ISA from typical cellulosic materials in the repository. Two different degradation experiments were conducted under alkaline conditions (saturated with Ca(OH)2 at pH 12.4): i) cellulosic material mixture under an opened condition (partially aerobic), and ii) cellulosic material under an anaerobic condition in a nitrogen-purged glove box. In the first case, three different types of cellulosic materials–paper, cotton, and wood– were mixed at the same ratio, and the experiments were carried out at three different temperatures (20°C, 40°C, and 60°C). It revealed that both the cellulose degradation rate and generated ISA concentration were high at high reaction temperatures, and various soluble degradation products such as formic acid and lactic acid were generated. The cellulose degradation in this work seems to still stay at a peeling-off process. In the second study, each type of cellulosic material was applied in its own batch experiments, and the amount of generated ISA was in the order of paper > wood > cotton. The above two experiments are supposed to be a long-term study until the generated ISA reaches an equilibrium state.

Recently, concern regarding disposal of cellulosic material is growing as cellulose is known to produce complexing agent, isosaccharinic acid (ISA), upon degradation. ISA could enhance mobility of some radionuclides, thus increasing the amount of radionuclide released into the environment. Thus, evaluation on the possible impact of the cellulose degradation would be an important aspect in safety evaluation. In this paper, safety assessments conducted in Sweden and UK are studied, and the factors required to be considered for appropriate safety assessment of cellulose is analyzed. SKB (Sweden) conducted safety assessment of cellulose degradation as a part of long-term safety assessment of SFR. SKB determined that ISA would impact sorption of trivalent and tetravalent radionuclides (Eu, Am, Th, Np, Pa, Pu, U, Tc, Zr and Nb) at concentration higher than 10−4–10−3 M, and impact sorption of divalent radionuclides (Ni, Co, Fe, Be and Pb) at concentration higher than 10−2 M. Then, SKB conservatively set the upper limit of ISA concentration to be 10−4 M and conducted cellulose degradation evaluation on each waste package type, considering the expected disposal environment of SFR. Based on the calculated results, some of the waste packages showed concentration of ISA to be higher than 10−4 M, so SKB conservatively developed waste acceptance criteria to prevent ISA being produced to an extent of affecting the safety of the repository. SKB conducted safety assessment only for the repositories with pH above 12.5 and excluded 1BLA from the safety assessment as the expected pH of 1BLA is around 12, which is insufficient for cellulose to degrade. However, SKB set disposal limit for 1BLA as well, to minimize potential impact in future. Serco (UK) conducted safety assessment of cellulose degradation for the conceptual repository, which is a concrete vault with cementitious backfill. Serco estimated that the pH of repository would maintain around 12.4. Serco conservatively assumed that the pH would be sufficient for cellulose degradation to occur partially, and suggested application of appropriate degradation ratio for safety assessment of cellulose degradation. To conduct appropriate safety assessment of cellulose degradation, an appropriate ISA concentration limit based on radionuclide inventory list, and an appropriate cellulose degradation ratio based on the pH of disposal environment should be determined. As for guidance, below pH 12.5, cellulose degradation is not expected, and between pH 12.5–13, partial cellulose degradation is expected. In future, this study could be used as fundamental data to evaluate safety of the repository.

Many basidiomycetes, especially mushroom species, thrive on saccharides that they obtain from the decomposition of cellulose, hemi-cellulose and or lignin. Cellulose is degraded by specific enzymes called cellulases. Cellulases play an important role in the biosphere by recycling cellulose, the most abundant carbohydrate produced by plants. Also cellulases have many potential biotechnologic and industrial applications. Detection of cellulose degrading activity is commonly performed on carboxymethylcellulose CMC medium in combination with a stain that reveals the degraded CMC. In order to efficiently screen the F. velutipes knockout library, obtained through Agrobacterium-mediated transformation (ATMT), for those important enzymes we compared two different staining methods i.e. Congo Red and Gram’s Iodine (as reported in R. C. Kasana et al 2008). The latter stain showed strongly enhanced detection in time and intensity, facilitating mass screening of our mutant database. Using Gram’s iodine and square petri dishes containing up to 9 colonies at a time we can now rapidly screen multiple mutants in a short period. We are going to find the genes. Inactivated genes of mutants with altered cellulase degradation activity will be identified and cloned for further analysis.

The genes encoding cellulases, which belong to glycosyl hydrolase families have been cloned from the basidiomycetous mushrooms. The transcripts of cellulase genes are strongly induced when the mycelia are grown in medium containing crystalline cellulose, and they are not expressed in medium containing glucose, but how insoluble substrates such as microcrystalline cellulose are recognized by these fungal cells is not clear. The polypore mushroom Polyporus arcularius is a wood-decomposing basidiomycete that produces at least three types (I, II, and IIIa) of carboxymethyl cellulase (CMCase) when the medium contains crystalline cellulose as the sole carbon source and produced mainly cellobiose in the medium. The genomic and cDNA clones encoding the family 12 endoglucanase (CMCase IIIa) gene (cel3A) of P. arcularius have been sequenced, and Cel3A has been expressed as an active enzyme in Escherichia coli. To determine the role and function of each type of cellulase in the degradation of crystalline cellulose by basidiomycetous mushrooms, the structure of all of the cellulase genes should be investigated, but the nucleotide sequences of the other cellulase genes in P. arcularius have not yet been reported. In the current study, the genomic and cDNA clones encoding the endoglucanases (cel4), and the two cellobiohydrolases (cel1 and cel2) of P. arcularius sequenced and characterized. The predicted amino acid sequence of Cel1 Cel2, Cel3a and Cel4 are similar to glycosyl hydrolase family 7, 6 12 and 5 protein, respectively. The expressions of the all cellulase genes (cel1 cel2, cel3a and cel4) were induced by Avicel (microcrystalline cellulose) and cellopentaose but repressed by glucose, cellobiose, cellotriose, and cellotetraose. There was a low level of transcription of both genes regardless of the carbon source. These results suggest that P. arcularius cells constitutively express a very low level of cellulase that can degrade insoluble crystalline cellulose and that the transcription of celluases in the cells is induced by products produced by these endoglucanases such as cellooligosaccharides. From our findings, we propose a possible mechanism for the recognition and degradation of insoluble crystalline cellulose by fungal cells.