Conservation Research: Fumigation and ... - ACS Publications

10 Conservation Research: Fumigation and

Downloaded by UNIV OF CALIFORNIA SANTA BARBARA on May 20, 2018 | https://pubs.acs.org Publication Date: December 17, 1978 | doi: 10.1021/ba-1977-0164.ch010

Sterilization of Flood-Contaminated Library, Office, Photographic, and Archival Materials D A V I D J. F I S C H E R Milton Roy Co., P.O. Box 1899, Sarasota, Fla. 33578

One of the consequences of tropical storm Agnes, June 1972, reached as far north as Corning, N.Y. The Corning Museum of Glass' library, the library of record for the world's glass scholars, sustained damage when flooded by muddy waters from an adjacent river. All of the library's damaged collection was frozen to prevent microbiological attack. A team developed a plan, which included special research projects, for the restoration of this collection. One of the major, successful projects was a process for not only fumigating but sterilizing the collection.

/^%n June 23, 1972 the city of Corning, N . Y . was hit by a devastating flood. The muddy waters rose to a height of five feet, four inches on the main floor of the Corning Museum of Glass. As the waters receded, a layer of m u d was deposited on the museum's priceless glass collection, its library of record, archives, slide and film collection, and office files. Because the m u d contained microbial contaminants as well as eggs from various insects, a l l of the flood-damaged materials except the glass objects were frozen to preserve them from attack by these biological species. This freezing action also permitted the museum's staff to con sider restoration plans specifically designed for each of these damaged items. E v e n though some of the museum holdings were not directly i n con tact with the flood waters, a l l became contaminated during subsequent removal from the museum and transfer to temporary storage areas. Since the humidity and temperature were not controlled i n these storage areas, there was serious concern about the possibility of biological growth i n 139 Williams; Preservation of Paper and Textiles of Historic and Artistic Value Advances in Chemistry; American Chemical Society: Washington, DC, 1978.

140

PRESERVATION OF PAPER

AND TEXTILES


these materials. A random check of the storage area, books i n the collec tion, and correspondence i n the office files substantiated this concern. Insect, fungal, and bacterial growth were found, and an immediate method to solve this problem was required. Starch, casein, and cellulosic materials, constituents of paper formulations, were being utilized by some of these species. Approach to Fumigation and Sterilization of Contaminated Books As restoration planning evolved, concern for the stored, nonfrozen contaminated materials increased. The use of insecticides was considered initially; however, the suggestion was eliminated after discussions were held with several groups of paper conservators. They felt that such com pounds might leave residual halogens i n the paper. Such halogens ulti mately contribute to an acidic environment which i n turn accelerates aging of the paper. This need to treat biological contamination necessitated the search for and purchase of appropriate equipment that could be operated by the museum staff. To minimize capital expenditures and as a result of the need for quick delivery and of the fact that only a small amount of space was available, a modified Vacudyne Fumigator ( V D F ) designed for use i n libraries was selected. It was manufactured with an extra-large vacuum pump and had heating elements installed i n its side walls. The unit was sold (Vacudyne Altair, Chicago Heights, 111.) as an instrument to establish conditions for k i l l of insects and mold, not as a sterilizer for k i l l of bacteria. Since ethylene oxide could be used i n its chamber, how ever, modification of its normal operating sequence was considered to develop a sterilizing treatment for the books. Ethylene oxide, when used under the proper conditions, is not only a fumigant but also an effective sterilizing agent. It is used extensively by the pharmaceutical industry to sterilize various plastics and bandages. Fumigating to destroy book worms, silverfish, moth, larvae, mold, and fungal contaminants proceeds under milder conditions than those required for killing the bacteriological contaminants. Since it was neces sary to sterilize a l l of the museum's books to eliminate subsequent dam age resulting from biological growth or transfer of contaminants by users to other books as well as to the users themselves, the following experi mental approach was developed. To obtain the maximum amount of information on the quality of sterilization with the least effort, attention was focused primarily on identifying conditions for k i l l of what was believed to be one of the most resistant bacterial species. After being procured, this species was to be placed i n the center of closed books, the location most difficult for gas penetration. If process conditions could be

Williams; Preservation of Paper and Textiles of Historic and Artistic Value Advances in Chemistry; American Chemical Society: Washington, DC, 1978.

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Fumigation and Sterilization of Flooded Materials

141

identified for k i l l at this location, the assumption that a l l other less resistant forms of biological life would also be killed was believed to be valid. Time and money would be saved by not conducting the latter experiments. A survey of the museum's problems confirmed that mold and bacterial growth were even more of a threat than insect growth.


Experimental Considerations Fumigation. The initial effectiveness of the V D F was determined by conducting a series of experiments which varied the amount of air being evacuated before the ethylene oxide mixture was allowed to enter, the time of exposure, and the temperature of application to the books. The dimensions of the chamber i n the fumigator were 1.2 m X 0.92 m X 0.46 m. A normal library cart with three shelves would be moved i n and out of the chamber. W h e n these shelves were stacked with books, approximately two-thirds of the volume inside the chamber was filled by the books and the cart. After loading the chamber and closing the door, a preselected program—evacuation of air, injection of gas for treat ment, evacuation of gas, and back-filling with air—was begun. Ethylene oxide at a composition of 12% by weight with U C O N refrigerant 12 (Oxyfume-12, Union Carbide Corp.) is nonflammable and can be used safely i n a closed system. The effectiveness of this commer cial mixture depends on the amount of ethylene oxide present, the tem perature of its application, the time of exposure, and the relative humidity in the chamber (1,2, 3). Microbiological Tests and Controls. Because Bacillus subtilis is a rugged, resistant microbiological species, spore strips (Castle C o . , Rochester, N . Y . ) of this bacteria were chosen for the study. The investi gation was supplemented by use of additional biological samples involv ing actual microbiologically attacked pages from badly damaged books; colonies of mold and bacteria had been allowed to mature intentionally for use i n these experiments. Later i n this paper, these samples which were placed inside books are described as muddy-moldy pages. Cultures of Escherichia coli and Aspergillus sp. were also used i n the experiments. Positioning of Biological Test Samples. Spore strips and sections of muddy-moldy pages were placed initially inside closed books. A l l sam ples were held at the center of the page with pressure sensitive tape. W h e n k i l l was not observed, the samples were again attached to a page in the book which was now set on its bottom edge and left partially opened during treatment. During the final experiment, when better conditions for k i l l were identified, special attention was given to the positioning of biological samples. They were attached to pages i n the center of books which were then placed between two 1.27-cm Plexiglas plates. Four C-clamps, one placed at each corner of the plates, were tightened with equivalent torques, 40 kg-cm, to ensure the uniform application of pressure to each book. This amount of torque was chosen to duplicate the pressure exerted on books when they are tightly packed on the library cart shelf. F i v e different locations on the cart, one at the outside edge of the top and bottom shelf and one i n the center, were used for the books containing biological samples.



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Trypticase soy agar and Sabouraud's agar, i n petri dishes, were streaked with E. coli and Aspergillus sp. and placed on top of books packed on the cart. The cart was placed i n the chamber and exposed to ethylene oxide for 4 hr. These petri dish samples were used for a special part of the second series of experiments. Microbiological Media. Trypticase soy broth ( l i q u i d ) , trypticase soy agar for bacteria, and Sabouraud's agar for fungi were used as media. Although the agar media are identical to and nutritionally comparable with the liquid media, the agar was added as a solidifying agent for cultivation of bacteria or fungi on a semisolid surface. Culturing of Test and Control Samples. Both control and treated spore strip samples were cultured at 37 °C i n trypticase soy broth for 48 hr to determine kill. Swab samples, taken from sections of the surface of the muddy-moldy pages before and after exposure to ethylene oxide, were placed i n 1 m L of 0.001M phosphate buffered saline solution at p H 7.0 for plating on appropriate media. These samples were taken from a 6.45-cm section. To determine the effect of adding moisture to dried contaminated pages, 1 m L of the phosphate buffered saline solution at p H 7.0 was added to a section of each of several muddy-moldy pages before treatment with ethylene oxide. Swabs were taken from these selected areas before and after the gaseous treatment. Duplicate plates were made of all samples. Process Conditions for Ethylene Oxide. Exposure to ethylene oxide lasted 4, 6.5, and 15 hr. To keep the relative humidity at the necessary percent ( I , 2, 3 ) , water vapor was added to the chamber of the V D F . Exposure of the books to ethylene oxide occurred at either room tem perature or at an elevated temperature. W h e n the maximum temperature in the chamber in the latter case was set for approximately 40 °C, it took about 60 min to get to 33 °C and 3 hr to reach the maxium value. The temperature changes of the gas throughout the chamber after closing the chamber door were determined by placing a series of thermocouples both inside and outside of the books at different positions on the cart. A maximum difference of 11 °C was recorded upon entry of the gas into the chamber between a pair of these thermocouples positioned on the bottom shelf. The difference was reduced to 1°C after 8 min of exposure. Temperatures at the various positions were monitored throughout the entire experiment. A comparable discrepancy was observed for the extreme i n the chambers gas temperature after entry of ethylene oxide. Temperature measurements were made between the gas occupying the volume just adjacent to the gas inlet (located at the bottom and rear of the chamber) and the volume i n the center of the chamber. The initial disparity in temperature as a result of injecting the gas was reduced to almost 4°C after 7 m i n of exposure. The evacuation level and the volume of the books and cart i n the chamber controlled the amount of ethylene oxide present in the chamber. The first series of tests was made when the air pressure i n the chamber was reduced to a value of 15 i n . H g . A t this pressure, a solenoid valve was opened to release the ethylene oxide mixture. Gas continued to enter until a pressure of 1 in. H g was attained; then the valve was shut. W h e n the second and third series of experiments were conducted, evacuation 2


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Fumigation and Sterilization of Flooded Materials143

in excess of 28 i n . H g took place before the solenoid valve was opened for entry of the ethylene oxide into the chamber. After the desired exposure time had elapsed, the chamber was evacuated to this pressure again and then filled with filtered air before it was opened.


Experimental Results and Discussion Series I. The initial series of experiments was conducted b y placing the spore strips inside closed books at five different locations on the cart and exposing for 4 hr and 14 hr at room temperature. The limit of the evacuation pressure was set at 15 i n . H g . Approximately 50 books were positioned on the cart, occupying about two thirds of the volume inside the chamber. None of the spores was killed by these treatments. F u r thermore, when the chamber was opened, the U C O N refrigerant 12 could be detected. These two results suggested that (1) an increase i n the total amount of ethylene oxide was required to facilitate k i l l and (2) the V D F program should be modified so that gas left after treating the books could be removed before the chamber door was opened. One change i n the V D F program satisfied both needs. The level of evacuation was adjusted to a pressure of 28 i n . H g before ethylene oxide entered the chamber and before the chamber was back-filled with air. Series II. I n this series of experiments both spore strips and muddymoldy pages were used, some placed i n books which were partially opened and set on their bottom edges while they were being treated. Moisture was added to some sections of the completely dried, contami nated pages. The results listed i n Table I show that spores of B. subtilis were not killed i n 15 hr when located inside closed books. W h e n a book was left partially open to permit ready access of ethylene oxide, all spores were killed.

Table I.

Treatment of Spore Strips at Room Temperature

Treatment Number of Time Individual (hr) Samples 0

Results after Incubation Location of Spore Strips

4

5

15

5

Inside closed books

15

5

Inside partially opened books, standing on edge

Inside closed books

Unexposed Exposed Strip Strip b

+ + +

+

+

° One at outside edge of top and bottom shelf and one at the center of the cart. Growth = + and no growth = —. 6


144


Table II.


Treatment Time (hr)

AND TEXTILES

Treatment of Muddy-Moldy Pages at Room Temperature Number of Individual Samples 5

a

5

a

Bacteria and Fungi Counts/6.1^5 cm

2

Location of Sample Inside closed book

Before TNTC

Inside partially opened book

Inside partially opened book

0

TNTC

After

Before

After

TNTC

TNTC

TNTC

TNTC

TNTC

TNTC

60

TNTC

TNTC

TNTC

Placed in open petri dish" 15

Fungi

Bacteria

One at outside edge of top and bottom shelf and one at the center of the cart. Located at outside edges of top shelf on cart. o T N T C = too numerous to count. Sample moistened with 1 m L phosphate buffer. a

h

d

The results described i n Table II suggest that a necessary minimum amount of moisture encouraged k i l l of fungi and bacteria at short expo sure times; k i l l was observed on the moistened muddy-moldy pages put i n the petri dishes on top of the books within 4 hr. If the pages were dry, a 15-hr exposure was required for k i l l of these same species. The latter findings, involving exposure of muddy-moldy pages taped to pages i n partially opened books, supported the data noted i n Table I for similar exposure and k i l l of spores. Table III. Treatment of Cultures Streaked on Nutrient at 4-hr Exposure

Species

Nutrient

Escherichia coli

Trypticase soy agar

Aspergillus sp. 1

Growth =

Sabrouraud's agar

+ and no growth =

Number of

Growth after Incubation 0

Samples Before After 5 5

+ +

—.


-


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145

A n additional experiment (see Table III) used streaks of moist vegetative cells of bacteria and moist fungal species placed on nutrients in petri dishes. Again it appears that only a 4-hr treatment at room temperature was required to k i l l these moist species. However, this con clusion assumes k i l l b y direct contact with the gaseous ethylene oxide as contrasted to subsequent k i l l because of ethylene oxide being absorbed by the agar and gradually released thereafter. Similar dehydrated froms (see Table I I ) , present on dry muddy-moldy pages, required 15 hr of exposure. The necessary moisture content and its location appear critical. The presence of water at a site other than biological could produce a reaction with ethylene oxide which would eliminate its chance to sterilize. Series III. During these tests, conditions comparable with the second series were maintained except that the chamber was heated. It is obvious from a review of Table I V that a 6.5-hr exposure is sufficient to k i l l the spores i n the most tightly packed books. There was a high percentage of k i l l of bacteria and fungi on the muddy-moldy pages (Table V ) placed in the same books and treated at the same time, but the k i l l was not total. These facts imply that longer exposure times are required for complete k i l l when contamination is excessive, m u d is present, and the surface of the page is dry. Complete k i l l was obtained when a 15-hr (Table V ) exposure was applied to muddy-moldy pages with populations of con tamination comparable with those treated during the 6.5-hr exposure at the same elevated temperature.

Table IV. Treatment and Evaluation of Spore Strips at Elevated Temperature Evaluation of Growth after Different Exposure Times Type of Book Storage Partially opened V

))

J)

V

Clamped shut

Position on Cart'

4hr Controls

0

6.5 hr Exposed —

+

— — — — —

+

+ 2 + 3 + 4 + 5 + 1 + 4 + + + and no growth = —. 1

Controls

+ + +

+

+

Exposed

15 hr Controls

+ + + + +

+ +

Exposed — —

— — —

— —

° Growth = 1, 2: edges of top shelf; 3: center of intermediate shelf; 4, 5: edges of bottom shelf. 6


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Table V . Evaluation of Treated Muddy-Moldy Pages at Elevated Temperatures


Type of Book Storage

Position on Cart

Expo- Bacteria and Fungi Counts/645 cm sure Fungi Bacteria Time (hr) Before After Before After

2

TNTC TNTC TNTC TNTC TNTC

0 0 0 0 0

TNTC TNTC

0 0

0 50 0

TNTC TNTC TNTC

0 0 0


0 70 0 10 50


20 10 10 30 10

6.5 6.5

TNTC TNTC

80 300

TNTC TNTC

60 0

6.5 6.5 6.5

TNTC TNTC TNTC

50 0 180

TNTC TNTC TNTC

0 10 10

15 15

TNTC TNTC

0 0

TNTC TNTC

0 0

15 15 15

TNTC TNTC TNTC

0 0 0

TNTC TNTC TNTC

0 0 0

Partially opened

1 2 3 4 5

4 4 4 4 4

TNTC" TNTC TNTC TNTC TNTC

Clamped shut

1 4

4 4

TNTC TNTC

Closed, lying on side

2 3 5

4 4 4

TNTC TNTC TNTC

Partially opened

1 2 3 4 5

6.5 6.5 6.5 6.5 6.5

Clamped shut

1 4


2 3 5

Clamped shut

1 4


2 3 5

0 0 0 0 0 TNTC TNTC

• TNTC = too numerous to count. During the Series III experiments, A T I Sterilization Indicator Tapes, #00180 (Aseptic-Thermo Indicator Co., North Hollywood, Calif.) were mounted beside a l l spore strips and muddy-moldy pages placed i n the books used for the 6.5- and 15-hr treatments. The color change from yellow to dark blue suggested that an environment for potential steriliza tion existed i n each case. The tapes could not guarantee micro-environ mental sterilization conditions when excessive growth had left biological debris or when m u d had been deposited. Such factors may control reac tions with ethylene oxide or limit its diffusion to the active biological site.


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Summary Sterilization of spore strips and muddy-moldy pages in books treated at room temperature was accomplished in 15 hr when the books were set on their edges and left partially open. The importance of sufficient mois ture is obvious from the results of treating the muddy-moldy page for 4 hr after being dampened. A convenient practical way to ensure the supply of the necessary amount of moisture to each biological site on each page still needs to be developed. If library books which had not been exposed to muddy water or had not experienced excessive bacterial and fungal growth are involved, a 6.5-hr treatment under the conditions listed in Series III is sufficient to sterilize even tightly closed books. Treatment of the most seriously con taminated books ( muddy-moldy pages ) required an excess of 6.5 hr and less than 15 hr of processing when the conditions outlined in the Series III investigations were met. The use of the higher vacuum, allowing a greater amount of ethylene oxide to enter the chamber, facilitated kill of the biological species. After treatment and back-filling with filtered air, traces of the U C O N refriger ant 12 gas were not detected upon opening the chamber door at the end of the sterilization sequence. The results from the above investigations with books led to the successful application of this process for sterilizing the library holdings, archives, office files, film, and photographic prints of the Corning Museum of Glass. Acknowledgments Robert E. Kephart, Professor of Microbiology at the Corning Com munity College, conducted all of the biological tests. The author is grateful for his direct assistance in planning and implementing these investigations. The author is also indebted to R. H . Brill and J. H . Martin of the Corning Museum of Glass for their encouragement of these inves tigations. Appreciation is due Robert M . Organ, Chief of Conservation, Analytical Laboratory at the Smithsonian Institute, for his information on uses of ethylene oxide. Part of the funds for conducting this research were obtained from the National Museum Act and the National Endowment for the Arts. Literature Cited 1. Ernst, R. R., Shull, J. J., "Ethylene Oxide Gaseous Sterilization—Concen tration and Temperature Effects" Appl. Microbiol. (1962) 10, 337-341.


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PRESERVATION OF P A P E R A N D TEXTILES

2. Gilbert, B. L., Gambill, F . M., et al., "Effect of Moisture on Ethylene Oxide Sterilization," Appl.Microbiol.(1964) 12, 496-503. 3. Opfell, J. B., Wang, Y., et al., "Penetration by Gases to Sterilize Interior Surfaces of Confined Spaces," Appl. Microbiol. (1964) 12, 27-31.


R E C E I V E D April 15, 1977.


Conservation Research: Fumigation and ... - ACS Publications

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