Blue-Biotechnology e la pulitura biologica di manufatti storico-artistici
10.6092/issn.1973-9494/7169
La ricerca di nuove strategie di pulitura è in costante crescita, trovando applicazione in molte procedure di restauro. Possiamo affermare che il ricorso a proteine enzimatiche, utilizzate in questi decenni, rappresenta un’innovazione nel campo della pulitura, poiché gli enzimi agiscono selettivamente sugli strati da rimuovere, senza incidere negativamente sui materiali costitutivi del manufatto. In questo studio si presentano i risultati preliminari della bio-rimozione del Paraloid B72 ricorrendo all’uso di un nuovo set di proteine enzimatiche, esterasi, estratte da organismi marini.
application/pdf
1973-9494
7169
en
Franco Palla
Conservation Science in Cultural Heritage
artwork restoration
Blue-Biotechnology and Biocleaning of Historic­-Artistic Artifacts
Text.Serial.Journal
https://conservation-science.unibo.it/
16
Articles
2017-07-24
2017-07-17
2017-07-24
2017-09-04
185-196
CSCH
Franco Palla
2016
10.6092/issn.1973-9494/7169
185
Università degli Studi di Palermo
1973-9494
artwork restoration
en
196
Wolbers, R.C., (2000) Cleaning Painted Surfaces: Aqueous Methods. London: Archetype Publications.
Blue-Biotechnology and Biocleaning of Historic­-Artistic Artifacts
https://conservation-science.unibo.it/article/download/7169/6897
16
B     LUE-BIOTECHNOLOGY       AND
      HISTORIC­-ARTISTIC ARTIFACTS
                                                               BIOCLEANING                    OF


   Franco Palla*6
   Laboratorio di Biologia e Biotecnologie per i Beni Culturali
   Dipartimento di Scienze e Tecnologie Biologiche Chimiche e Farmaceutiche
   Palermo, Italy

   Keywords: bioactive molecules, esterase, Paraloid-B72, artwork restoration


   1. Introduction

   Cleaning is frequently one of the first actions in a restoration project. However, it can
be seen as going against the principles of conservative restoration as it is an irrevers-
ible procedure and an action that may produce a visible change on the artwork surface.
Cleaning is undoubtedly able to re-establish the correct reading of a work of art (Figure
1) and is particularly justified if its benefits and the chosen procedure are compatible
with the constituent materials and provide the lowest possible risk level for the artifact
itself and for restorers’ health.




                                                                                                            CONSERVATION SCIENCE IN CULTURAL HERITAGE




Figure 1. Cleaning, clearly showing re-establishment of the correct reading of the painting surface
(by E. La Francesca, Degree in Conservation and Restoration of Cultural Heritage, University of
Palermo, Italy).


* Corresponding author: franco.palla@unipa.it


                                                                                                      185
                                                                                    The ongoing search for novel strategies continues, others are in the process of be-
                                                                                 ing developed, but known strategies are the most appropriate in all restoration proce-
                                                                                 dures. One innovation in cleaning procedures is represented by enzymatic cleaning.
                                                                                 Since its action is highly selective, it effectively removes the undesired layers without
                                                                                 affecting the constituent or surrounding materials. Proteins with enzymatic activity are
                                                                                 hydrolases, which have been tested since the 1970s to remove starch paste, animal
                                                                                 glues or protein adhesives and aged acrylic coatings by using purified enzymes as
                                                                                 single or mixed solutions [1-4]. Enzymes, such as trypsin, amylase and protease
                                                                                 have been used, mainly for the treatment of glue layers on paper, but applications
                                                                                 have already been made on a variety of artworks, such as paintings on canvas, mural
                                                                                 paintings and wood and stone artworks. Commercial hydrolases, such as proteases,
                                                                                 amylases, lipases and esterase are isolated from animal (pancreas, stomach), veg-
                                                                                 etal (oat and wheat seeds) and microbial (bacteria or fungi) sources and utilized in
                                                                                 biocleaning treatments in order to remove specific substrates [5], also in combination
                                                                                 with the action of viable bacterial cells [6]. These methods are a very helpful tool in the
                                                                                 biocleaning of artworks, often representing a valid alternative to chemical solvents,
                                                                                 characterized by selective removal action and safety for both the restorer and the
                                                                                 environment [7].
                                                                                    A complete characterization of the surface layer/s to be removed can be performed
F. Palla - Blue-biotechnology and biocleaning of historic-artistic artifacts




                                                                                 by Fourier Transform Infrared (FTIR) Spectroscopy [8] or by using biochemical meth-
                                                                                 ods such as High Performance Liquid Chromatography (HPLC) [9] or Sodium Dodecyl
                                                                                 Sulphate-Polyacrylamide gel (SDS-PAGE) electrophoresis [10].



                                                                                    2. Novel hydrolases (Protease, Esterase)

                                                                                     In the last three years, promising results have been obtained from the collaboration
                                                                                 between the Biology and Biotechnology for Cultural Heritage and Immunology labora-
                                                                                 tories at the University of Palermo. Particularly, cold-active molecules, extracted from
                                                                                 marine invertebrate organisms have been utilized in restoration procedures to remove
                                                                                 protein layers. Preliminarily, laboratory tests were performed on ad hoc specimens,
                                                                                 and only after finding that our hypotheses were established, were the same tests car-
                                                                                 ried out on several artifacts. In a first step, bioactive molecules with protease activity
                                                                                 were applied in order to remove protein layers from a wooden artifact, an oil on canvas
                                                                                 painting, a wax sculpture [11-14]. Removal efficiency was evaluated by detecting the
                                                                                 molecules present on the cotton swabs utilized in the final cleaning procedure, by
                                                                                 Amido Black Staining Solution [15].
                                                                                     The research focalised on different undesired layers, mainly esters and waxes fre-
                                                                                 quently used in the restoration procedure [16]. Particularly, a set of insect (bee) or
                                                                                 vegetal (carnauba) waxes were selected and several related laboratory specimens
                                                                                 were assembled; at present the specimens are undergoing a process of artificial age-
                                                                                 ing (repeated cycles: UV-A 300–400 nm; T = 22 ± 5°C; RH = 60-65%).
                                                                                     In order to set up biocleaning procedures for the removal of these waxes a new
                                                                                 set of bioactive molecules was isolated from marine organisms, specifically BME
                                                                                 (bioactive molecules with esterase activity). In parallel, we also carried out a clean-
                                                                                 ing test on Paraloid B72 aged layers, a thermoplastic resin extensively utilized in the
                                                                                 field of artwork restoration as adhesive, consolidating agent, filler, etc [17]. In this
                                                                                 study, removal of the Paraloid B72 layer was performed on a laboratory specimen


                                                                           186
mimicking the “strappo” ‘of mosaic tesserae, carried out by gluing tesserae on a
canvas sheet; this specimen was naturally aged for 5 years (Temp=24±5°C and RU
= 58±10%).
   Interesting results were achieved after 30 minutes application of bioactive mole-
cules with esterase activity (BME) solution, at 26°C (laboratory environment tempera-
ture), as shown in Figure 2.




                                                                                                       CONSERVATION SCIENCE IN CULTURAL HERITAGE




Figure 2. Bio-removal of canvas layer stuck on mosaic tesserae by Paraloid B72: a) after 30
min of application of BME enzyme solution, the canvas layer is removed, but Paraloid residues
are still present on the tesserae; b) supplementary application for 10 min of enzyme solution
allows complete removal of resin from tesserae surface; c) UV-induced fluorescence support the
observation in b).



   3. Conclusion

    Enzymes are usually utilized in aqueous systems, consequently, they cannot be
utilized in all restoration procedures, but can, however, be applied as gelled solutions.
Several gelling agents (Klucel-G, Pluronic F108, Vanzan NFC 2) can be utilized as


                                                                                                 187
                                                                                 enzyme support (specific viscosity, water release), which is able to guarantee stable
                                                                                 reaction conditions and facilitate the cleaning procedure [13]. Moreover, the enzyme
                                                                                 action may be influenced by some inhibitors, found in a large number of ancient and
                                                                                 modern pigments, such as heavy metal ions (Cu2+, Pb2+, Cd2+, Hg2+, Sn2+), or by specific
                                                                                 pH and temperature values and application time.
                                                                                     Great progress has recently been made in the application of bioactive molecules
                                                                                 isolated from marine organisms such as sponges, jellyfish, sea-anemones, shellfish
                                                                                 (Blue-Biotechnology) representing an important resource useful to the health, food
                                                                                 and processing, and preservation industries. The peculiarities of these molecules are
                                                                                 stability, activity at low-temperature (<30°C) and specificity of action. In the field of
                                                                                 conservative restoration, the new hydrolase with protease (BMP) or esterase (BME)
                                                                                 activity tested in our studies showed selective action avoiding any damage to the con-
                                                                                 stitutive materials. Another peculiarity is the absence of any negative impact on restor-
                                                                                 ers and environment, thus falling into the category of green restoration technologies,
                                                                                 indispensable for the sustainable conservation of cultural assets.
                                                                                     Despite the evident positive aspects of the innovative procedure for the biologi-
                                                                                 cal cleaning of historic-artistic artifacts, a systematic study of how hydrolytic enzymes
                                                                                 work on artworks remains to be implemented, with a focus on optimizing the param-
                                                                                 eters that are crucial for catalytic activity compared to the nature of the undesired lay-
F. Palla - Blue-biotechnology and biocleaning of historic-artistic artifacts




                                                                                 ers that must be removed.


                                                                                       Acknowledgements

                                                                                     The author is indebted to the restorers, Mauro Sebastianelli and Eduardo La Franc-
                                                                                 esca, of the Diocesan Museum of Palermo, for their collaboration and all coworkers
                                                                                 from the Laboratory of Biology and Biotechnology for Cultural Heritage at the Univer-
                                                                                 sity of Palermo.


                                                                                       References

                                                                                 [1]     Wendelbo, O. (1976) The Use of Proteolytic Enzymes in the Restoration of Paper
                                                                                         and Papyrus. University Library of Bergen
                                                                                 [2]     Bellucci, R., Cremonesi, P., Pignagnoli, G. (1999) A note on the use of enzymes
                                                                                         in conservation: the removal of aged acrylic resin coatings with Lipase, Studies
                                                                                         in Conservation, 44(4), pp. 278-281.
                                                                                 [3]     Wolbers, R.C., (2000) Cleaning Painted Surfaces: Aqueous Methods. London:
                                                                                         Archetype Publications.
                                                                                 [4]     Segal, J., Cooper, D. (1977) The use of enzymes to release adhesives, Paper
                                                                                         Conservator, 2, pp. 47-50.
                                                                                 [5]     Palla, F. (2013) Bioactive molecules: innovative contribution of biotechnology
                                                                                         to the restoration of cultural heritage, Journal Conservation Science in Cultural
                                                                                         Heritage 13, pp. 369-373.
                                                                                 [6]     Ranalli, G., Alfano, G., Belli, C., Lustrato, G., Colombini, M.P., Bonaduce, I., Za-
                                                                                         nardini, E., Abbruscato, P., Cappitelli, F., Sorlini, C. (2005) Biotechnology applied
                                                                                         to cultural heritage: biorestoration of frescoes using viable bacterial cells and
                                                                                         enzymes. J Appl Microbiol, 98(1), pp. 73-83.


                                                                           188
[7]     Bosch-Roig, P., Ranalli, G. (2014) The safety of biocleaning technologies for
        cultural heritage. Front. Microbiol. 5, pp.1-3.
[8]     Sarmiento, A., Pérez-Alonso, M., Olivares, M., Castro, K., Martinez-Arkarazo, I.,
        Fernández, LA., Madariaga, JM. (2011) Classification and identification of organ-
        ic binding media in artworks by means of Fourier transform infrared spectroscopy
        and principal component analysis, Analytical and Bioanalytical Chemistry, 399
        (10), pp. 3601-3611.
[9]     Palla, F., Cammarata, M., Trapani, M.R., Salamone, M., Ghersi, G., Sebastianel-
        li, M. (2012) Novel protease from marine organisms with potential interest in the
        restoration procedure. In: Proceedings of International Congress on Science and
        Technology for the Conservation of Cultural Heritage, 2nd-5th October, Santiago
        de Compostela, Spain, pp. 279-282.
[10]    Laemmli, U.K. (1970) Cleavage of Structural Proteins during the Assembly of the
        Head of Bacteriophage T4, Nature, 227, pp. 680-685.
[11]    Barresi, G., Di Carlo, E., Trapani, M.R., Parisi, M.G., Chillè, C., Mulè, M.F., Cam-
        marata, M., Palla, F. (2015) Marine organisms as source of bioactive molecules
        applied in restoration project. Heritage Science, 3, pp. 17-20.
[12]    Chillè, C., Di Carlo, E., Barresi, G., Sottile, S., Lazzara, G., Cammarata, M.,
        Palla, F. (2014) Pulitura mediante proteine enzimatiche sperimentali. In: Pro-
        ceedings of XII Congresso Nazionale IGIIC – Lo Stato dell’Arte – Accademia Di
        Belle Arti di Brera – Milano 23/24 ottobre, pp.1-8
[13]    Palla, F., Ferrara, G., Richiusa, A., Teri, R.M.C., Sebastianelli, M. (2015) Applica-
        tion of new proteases in removal protein layers from artistic works surfaces. In:
        Proceedings of 7th ESRARC - European Symposium on Religious Art. Restora-
        tion & Conservation, Trnava 4-6 June, pp. 66-69.
[14]    Palla, F., Barresi, G., Giordano, A., Schiavone, S., Trapani, M.R., Rotolo, V.,
        Parisi, M.G., Cammarata, M. (2016) Cold-active molecules for a sustainable
        preservation and restoration of historic-artistic manufacts. International Journal




                                                                                                      CONSERVATION SCIENCE IN CULTURAL HERITAGE
        of Conservation Science, 7(1), pp. 239-246.
[15]    Martin, E. (1977) Some improvements in techniques of analysis of paint media.
        Studies in Conservation, 22, pp. 63-67.
[16]    Amoroso, G.G. (2002) Trattato di scienza della conservazione dei monumenti:
        etica della conservazione, degrado dei monumenti, interventi conservativi, con-
        solidanti e protettivi. Alinea Editrice.
[17]    Podany, J., Garland, K.M., Freeman, W.R., Rogers, J. (2001) Paraloid B-72 as a
        Structural Adhesive and as a Barrier within Structural Adhesive Bonds: Evalua-
        tions of Strength and Reversibility, Journal of the American Institute for Conser-
        vation, 40(1), pp. 15-33.



      Biographical notes

   Franco Palla is Associate Professor of Applied and Environmental Botany at the
University of Palermo, Italy. He is Professor and Coordinator of the Five-Year Degree
in Conservation and Restoration of Cultural Property (MRL02, certified professional
restorer). He is a member of the Permanent Commission for the Protection of Cultural
Heritage of the National Order of Biologists. Scientific head of UNIPA Research Unit,
for the Research Project PON01_00625, It@cha (Italian Technology for Advanced Ap-


                                                                                                189
                                                                                 plications in Cultural Assets). He was one of the members of the working groups in the
                                                                                 cooperation project Italy-Cambodia for Training Experts of Cultural Heritage, Universi-
                                                                                 ty of Palermo – Royal University of Fine Arts and the Ministry of Culture and Fine Arts,
                                                                                 Angkor, Cambodia. He is Coordinator of the Laboratory of Biology and Biotechnologies
                                                                                 for Cultural Heritage at the Department of Biological, Chemical and Pharmaceutical
                                                                                 Sciences and Technologies (STEBICEF) of the University of Palermo.
F. Palla - Blue-biotechnology and biocleaning of historic-artistic artifacts




                                                                           190
