APPENDIX

Variations to TAPPI Standard T487 om-93

The standard describes two methods, the “spore-mycelia” and the “cloud of spores.” Only the spore-mycelia method was used. Modifications consist of:

1. The form of the filter paper used for the samples was different from that indicated in the standard. In place of squares of 50 mm2, octagons of 420 mm2 were used. After the polymer coatings dried, each octagon was separated into four sections, each with an area of 105 mm2.

2. Fungi of a species recommended by TAPPI standard T487 om-93 were employed for test organisms, but not the exact strains because the exact ones were not obtainable.

3. Incubation period for the inoculated samples was 21 days instead of 14 days, at 28�C and 100% RH.

4. In place of observing presence or absence of filaments or colonies of fungi, evaluation of results was in values of zero-span tensile strength (newtons). Then, after drying, the samples were cut in strips 1.5 cm wide (see TAPPI 1985).

Percentage of Weight Loss

Five g of Whatman no. 1 filter paper was shredded in aseptic conditions. The shredded material was placed in a blender with the blades protected by duct tape to prevent fiber degradation. Enough deionized water was added to obtain a fiber slurry. This substance was then transferred to a glass filter, and water was removed by suction. The pulp was lifted out with a spatula, homogenized, and bagged and sealed in polyethylene. Moisture content was measured in a thermal balance.

For the next step, 5 g (oven-dry basis) samples were weighed and placed in 50 ml wide-mouthed glass jars, capped, and sterilized in an autoclave at 10.42 kilopascals for 15 minutes.

Strain Culture

Each strain was inoculated in a culture test tube with potato-dextrose-agar medium and was incubated 21 days until sporulation.

To inoculate the paper samples, each strain was prepared as follows. First, glass jars containing 100 ml of distilled water and 30 glass beads were autoclaved. Then, to each test tube, 5 ml of sterile water was added, and the mycelium was scraped with a wire loop, followed by vigorous shaking to produce a fragmented mycelium and spore slurry. The contents were then poured into the glass jars of sterile distilled water and mixed thoroughly.

This process was repeated for each strain. Later, 1 ml of the slurry was taken up with a sterile plastic syringe and applied to the paper pulp previously stored in the glass jars. The jars were immediately capped and incubated at 28�C for 28 days in a sealed plastic box containing a water layer in the bottom as a reservoir to keep relative humidity at 100%, monitored by a hygrometer.

After this time period, 70% ethyl alcohol was added to kill the fungi, then the sample was dispersed using a mixer with distilled water. It was then filtered through a glass filter with suction, washed with 70% ethyl alcohol, placed in small aluminum dishes and air-dried, and then oven-dried at 105�C until reaching constant weight. Results are reported in percentage of weight loss according to the following formula:

Degree of Polymerization

The paper samples and inoculation strains were prepared in the same manner as the tests for percentage of weight loss. Evaluating degree of polymerization was in accordance with ASTM standard method D4 243-99 (1999).

To do the viscosity determination, each sample was dissolved in a cupriethylenediamine solution, and the fluid velocity was measured in a viscometer. Ten replicates of each sample were made, and the averages were calculated. Results of the selected strains are shown in figure 6.

ACKNOWLEDGEMENTS

We wish to thank the following persons for their support and for making this work possible: Karl Augustin Grellman Institute of Wood, Cellulose and Paper, University of Guadalajara, Guadalajara, Jalisco: Jos� Turrado Saucedo, chemical engineer and head of the technology laboratory; Jos� de Jes�s Rivera Prado, chemical engineer, the bleaching laboratory; Salvador P�rez Ramos, chemical engineer, the physical-mechanical tests laboratory; Francisco Vel�zquez Cervantes, chemical pharmacology biologist, and Leticia Maya, of the chemistry laboratory; Professor Bruno Becerra Aguilar of the technology area; Hilda Palacios Ju�rez, chemical engineer, the microscopy laboratory; and Luz Elena Arce Castillo, chemical engineer, the library and publications area. Biologist Rosario V�squez of the Mycology Laboratory of Polytechnic National Institute in Mexico City. University Center of Biological and Agronomical Sciences, University of Guadalajara: Professor Rosa Mar�a Dom�nguez Arias. Mexican Institute of Social Security Clinic 46 in Guadalajara: Maria Luisa Preciado Quiroz and Mar�a de Jes�s Najar. Mercedes G�mez Urquiza, director of the Manuel del Castillo Negrete National School of Conservation, Restoration and Museology; Luis Daniel Mario Goeritz Rodriguez, engineer and director of the National Institute of Anthropology and History Veracruz Center. We also thank Daniel McGonagle for assistance in translating part of the text.

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SOURCES OF MATERIALS

Whatman no. 1 Catalog number H-06648-17 Equipar, S.A. de C.V. Juan S�nchez Azcona No. 1447 Colonia del Valle, delegaci�n Benito Ju�rez Mexico, D.F. C.P. 03100

Methocel F4M Premium CMC and MC Droguer�a Cosmopolita S.A. de C.V. Av. Revoluci�n No. 1080 Col. Mixcoax Mexico City, Mexico

The chitosan was prepared from shrimp shells by the Chemical Laboratory of the Karl Augustin Grellman Institute of Wood, Cellulose, and Paper, University of Guadalajara. Autopista Guadalajara-Nogales, kil�metro 15.5 Las Agujas Municipio de Zapopan Jalisco, M�xico Apartado Postal 52-93, C.P. 45020 Zapopan, Jalisco, Mexico

RK Print-Coat Instruments Ltd. Royston Herts 5G8 OQZ U.K.

Statgraphics Statistical Graphics System Educational Institution Edition—Version 7 Manugistics Inc. and Statistical Graphics Corporation 9715 Key West Ave. Rockville, Md. 20850

Sterilization with ethylene oxide was done in regional hospital number 46 of the Social Security Institute (IMSS) Calzada L�zaro C�rdenas y 8 de Julio Zona Industrial C.P. 44940 Guadalajara, Jalisco, Mexico

AUTHOR INFORMATION

MARIA DEL PILAR PONCE-JIM�NEZ received her biology degree from the National Autonomic University of Mexico and her master's degree from the University of Guadalajara with a specialty in cellulose and paper. Since 1990, she has been professor of conservation and restoration of paper and graphic documents at the National School of Conservation and Restoration and Museography at the Manuel del Castillo Negrete Institute of the National Institute of Anthropology and History (INAH) in Veracruz, Mexico. She also develops methods of teaching, application, and investigation in the field of paper conservation. Address: Centro INAH Veracruz, Benito Ju�rez No. 425 y 431, Zona Centro, C.P. 91700, Veracruz, Veracruz, Mexico.

FERNANDO A. L�PEZ-DELLAMARY TORAL received his B.S. from the National Autonomic University of Mexico and his Ph.D. from the University of Wisconsin at Madison. He is a professor of chemistry and analytical chemistry at the University of Guadalajara. He is chief of the chemistry laboratory at the University of Guadalajara's Karl Augustin Grellman Institute of Wood, Cellulose, and Paper, where he heads investigative projects on paper technology, biochemistry, and forestry engineering. Address: Departamento de Madera, Celulosa y Papel Karl Augustin Grellman, Apartado Postal 52-93, C.P. 45020, Zapopan, Jalisco, Mexico.

HUMBERTO GUTIERREZ-PULIDO holds a master's degree in statistics with a specialty in industrial productivity. He is a professor of design and experiment analysis in the advanced courses of the Exact Sciences and Engineering University Center at the University of Guadalajara. He does counseling, teaching, research, and applied projects in the field of statistics. Address: Centro Universitario de Ciencias Exactas e Ingenier�as, Boulevard General Marcelino Barrag�n y Calzada Ol�mpica, S. R. C. P. 44420, Guadalajara, Jalisco, Mexico.