The influence of substrate specificity of protease and other washing conditions on washing effect have been studied by using neck bands of cotton cloth soiled with skin grime, containing protein of 6-10 mg/g of cotton cloth, prepared by 20-30 persons for 1-3 days.
1) Considering the influence of substrate specificity of protease on the washing efficiency of soiled neck bands, the removal efficiency was rather low comparing to removal of stains caused by other proteins. However, when alkaline protease produced by Bacillus subtilisin BPN' and Streptomyces fradiae showed the removal efficiency was remarkably high owing to hydrolysis of skin grime.
2) With regard to the influence of protease specific activation on the effect of washing of soiled neck bands, the highest removal efficiency was observed when a solution of 1-3 PU/ml was used. Especially when alkaline protease which is stable in detergent solutions was used, the additive action of the detergent and protease resulted in a high removal efficiency.
3) With regard to the influence of the pH of the detergent solution on the effect of washing of soiled neck bands, the pH specificity of protease was reflected on the removal efficiency, and the optimum pH produced the highest removal efficiency.
4) With regard to the influence of the temperature of the detergent solution on the effect of washing of soiled neck bands, the decomposition and dispersion of the keratin layer was most readily affected at temperatures between 40°C and 60°C, resulting in a high removal efficiency. When alkaline protease produced by Stret'tomvices fradiae was used, the removal efficiency was remarkably high.

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The effect of washing on the number of bacteria removed from contaminated diapers was examined by using small pieces of cloth which were (1) either moistened with the water in which Proteus vulgaris OX19 was suspended or (2) spread with feces. The sample fabrics (1) were directly washed in a shaker, whereas the sample fabrics (2) were sewn onto larger pieces of cotton cloth and were washed by hand. The washing procedure was as follows : first, the fabrics were soaked into water for 30 minutes without stirring, then washed for 5 minutes, and were rinsed twice, each for 3 minutes. The above procedure was conducted (a) with detergent and (b) without detergent.
The removing effect of washing of the fabrics (1) is expressed by the rate of bacteria remaining unremoved on them (percentage of remaining number of bacteria to that of bacteria before washing) . The removal rate was : (a) 30% and (b) 50% after dipping, (a) 7% and (b ) 11% after shaking, (a) 0.4% and (b) 1.5% after two times of rinsings.
The rate of the destruction of bacteria through the entire process was 48% with the detergent group and 15% with the water (without detergent) group. Concerning the number of bacteria which remained unremoved on the feces-contaminated pieces after manual washing, the ratio of the fabrics washed with detergent to those washed without detergent was about 1 : 2. The degree of the unremoved yellow tincture caused by feces had a correlation with the number of unremoved bacteria.

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