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Biological Effects of Proteolytic Enzyme Detergents1

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Biological Effects of Proteolytic Enzyme Detergents1 Thorax: first published as 10.1136/thx.31.6.621 on 1 December 1976. Downloaded from Thorax (1976), 31, 621. Biological effects of proteolytic enzyme detergents1 A potentially serious respiratory hazard during later. A majority of the work presented was the manufacture of enzyme detergents was first unpublished. reported by Flindt (1969) and Pepys et al. (1969). In the eight years since the hazard was identified much effort has been put into investigating the PRODUCTION AND DUST CONTROL extent and nature of the effects and removing Van Veizen (Gist-Brocades, Netherlands) des- their cause. A feature of the work has been the cribed the manufacture of the enzyme by a fer- national and international exchange of medical mentation process, using Bacillus subtilis. The and industrial hygiene information between pro- purification steps need extremely careful quality ducing companies and the help provided by control. This is achieved by a batch process of academic departments of immunology and respira- manufacture. After purification (germ removal), tory disease to assess the biological effects. These a concentrated enzyme powder is produced by have included serial observations to monitor the precipitation, filtration, and drying. To ensure a efficacy of the rapidly improving dust conditions clean environment, the relatively dusty powder is achieved by the engineers. handled in a closed system. As in the detergent In 1974 Professor Hans Weill, of Tulane Uni- industry, great attention is now paid to engineer- versity, New Orleans, and Dr. John C. Gilson, ing detail where spillages or emissions of dust may Director of the MRC Pneumoconiosis Unit, sug- occur; for example, an ingenious pneumatic copyright. gested that 1976 would be an opportune time to double seal is used in the bagging of the powder. review the scene to obtain a general picture of In the detergent factory the heat-sensitive en- progress and of the lessons learnt which might zyme has to be incorporated in a powder base have applications elsewhere. The opportunity which is manufactured by spray drying in a large given by such a meeting was welcomed by the tower at elevated temperature. The manufacture companies in the USA and in Europe making the of the powder base was described by Davies enzymes and the enzyme detergent products. (Lever Brothers, UK) to give an indication of the http://thorax.bmj.com/ Topics covered included: making the enzymes scale of the manufacturing process (tons per and the detergents; toxicological and immuno- hour). After addition of the heat-sensitive en- logical studies in animals; clinical features of the zymes and other chemicals, the product is mixed effects in small groups and in individuals; immuno- and then put into cartons in the packing room. logical surveillance of exposed working groups; This was previously a labour-intensive area where and cross-sectional and longitudinal studies of exposures readily occurred. Great attention has respiratory symptoms and lung function of pro- been and is now continuing to be paid to reducing duction workers in several countries. There was these exposures. The dustiness of the product has on October 2, 2021 by guest. Protected an excellent opportunity, therefore, for engineers, been reduced by 'encapsulating' the enzymes. It epidemiologists, immunologists, occupational was glued to a coarse (>150 ,u) granule of phos- physicians, respiratory physiologists, and statisti- phate base by a tacky non-ionic detergent (an cians to contribute to the general assessment of ethoxylated linear aliphatic alcohol). The enzyme current knowledge in this field. Papers prepared manufacturers had now made further improve- for the meeting will be appearing in appropriate ments by forming 'marumes' and 'prills'. These journals. A list of the papers precirculated is given are beads containing the enzyme embedded in (p. 631) to assist in tracing the published articles non-dusty matrix. The 'prill' is produced by spraying a molten mixture of the enzyme with an 'Report of a symposium held on 4-5 May 1976 at the MRC organic material (non-ionic) into a tower in which Pneumoconiosis Unnit, Cardiff, sponsored by the Medical Research Council, UK, and the Soap and Detergent Industries Association the droplets cool to form regularly shaped non- (SDIA), UK. For the list of participants with their affiliations and the titles of papers presented, see p. 630. The report was prepared by J. C. dusty solid beads. By this means the enzyme is Gilson, C. P. Juniper, R. B. Martin, and H. Weill with the assistance incorporated into a less friable matrix than in the of Rapporteurs-D. R. Davies, J. H. Edwards, C. P. Juniper, W. E. Parish, and C. E. Rossiter. earlier methods of encapsulation. 621 Thorax: first published as 10.1136/thx.31.6.621 on 1 December 1976. Downloaded from 622 Biological effects of proteolytic enzyme detergents Bruce (Procter & Gamble, UK) described the much less marked. The haemorrhagic response in- advances in dust monitoring. The SDIA had duced by high concentration of enzymes could developed a high-volume (700 1 /min) Galley also be demonstrated on the exposed vascular sampler with a performance similar to the MSA bed of the rat cremaster muscle preparation. Heat- Fixt Flo. They also used a Casella personal ing the enzyme preparation prevented pulmonary sampler (2 1/min). At low dust concentrations and cremaster muscle injury. the instruments agreed well; at high levels In the guinea-pig repeated weekly exposures to (>02 Gu/m3; Gu=Glycine units, see Glossary aerosols at a level which did not induce pul- of Terms, p. 628) the personal sampler gave values monary damage at a single exposure resulted in of up to 10-fold greater. The reasons for this were systemic sensitization after the third or fourth not yet clear. A continuous sampler was devel- exposure. Mediation of the immune system was oped; this is useful in detecting peak emissions of shown by an immediate skin reaction, positive enzyme. The use of an Anderson sampler had Schultz-Dale reaction, and passive cutaneous shown that about 50% of the dust collected was anaphylaxis reactions (PCA). The latter reaction respirable (<7,u), and 70% of the enzyme dust established that the predominant circulating anti- was in this respirable fraction. However, there body was IgGla. With continuing exposures at was some difficulty in interpreting the results of weekly intervals, the respiratory response de- an Anderson sampler with a dust consisting of creased and was virtually absent after 20 ex- fragile particles which might be fractured into posures. This was associated with an increase in smaller pieces during passage through the the total level of specific enzyme inhibitor in the instrument. serum. Davies used the results of dust assessments from The difference in responses of isolated normal one factory from 1969 (when the first measure- or 'sensitized' guinea-pig lungs, when challenged ments were made) to 1975 to show the secular with enzyme preparations, confirmed that the trend of improvement achieved by following the induced in normal lung was bronchospasm copyright. SDIA recommendations (SDIA, 1969). During the associated with the release of histamine, whereas six years there had been a six-fold reduction in the effect on sensitized lung was associated with the average atmospheric dust concentration in a greater release of histamine and SRS-A. In vivo the packing rooms of the four UK detergent fac- studies using specific antagonists confirmed these tories (1200-*200 tLg/m3), and in the same period findings. the amount of associated enzyme dust had been The effect of detergents on the immune re- reduced by a factor in excess of 30 (2-3 Gu/m3) sponse was seen only at high detergent levels. http://thorax.bmj.com/ in 1969 to 0 05-0l1 Gu/m3 in 1975. Further When guinea-pigs were exposed to very high levels improvements had been achieved by the use of of a detergent aerosol (anionic or non-ionic; 'marumes' and 'prills' (SDIA Medical Review 70 mg/m3) in the presence of an enzyme aerosol, 1969-75). Atmospheric enzyme levels were now which alone did not produce sensitization at the present limits of analytical detectability. (6000 Gu/m3), a 40% incidence of sensitization occurred. Exposure for a total of 3500 hours TOXICOLOGY (18 h/day, 4 days/week, for 1 year) to dust eluted from enzyme washing powder product, at levels Cambridge (Unilever, UK), described the effects on October 2, 2021 by guest. Protected of heavy exposures in guinea-pigs and rats to of about 2 mg/m3, with enzyme activity 1 Gu/m3, aerosols of solutions of enzyme preparations. produced sensitization in only 1 of 16 guinea- The effects were dose-related. Guinea-pigs ex- pigs. posed to 80 000 Gu/m3 for one hour (enzyme Ritz (Procter & Gamble, USA) reported on activity 1100 Gu/mg) showed respiratory distress the relative anaphylactogenicity and immuno- and hypothermia with some fatalities. Histologi- genicity of a number of commercial enzyme pre- cally, intra-alveolar haemorrhage with eosinophil parations-Alcalase, Rapidase, and Monsanto and neutrophil infiltration was seen. At a quarter DA-lO-together with an experimental Alkaline of the dose for half the time only eosinophil- Protease. Guinea-pigs were given enzyme intra- neutrophil infiltration was seen. Similar effects tracheally weekly or biweekly in the range of were observed in animals exposed to dry enzyme 10 ng-10 ,ag (protein nitrogen) per dose. The powder. The rat was much more resistant to the pulmonary response was graded in severity on a effect of exposure to enzyme aerosols; even at the five-point scale. Serum samples were examined by highest concentration alveolar haemorrhage was PCA for the presence of IgGl and by immuno- Thorax: first published as 10.1136/thx.31.6.621 on 1 December 1976. Downloaded from Biological effects of proteolytic enzyme detergents 623 diffusion for precipitating antibodies against the greater than that of human, rabbit, or horse lung, enzyme.
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