Wall paintings are integral to cultural heritage and offer rich insights into historical and religious beliefs. There exist various wall painting techniques that pose challenges in binder and pigment identification, especially in the case of egg/oil-based binders. GC-MS identification of lipidic binders relies routinely on parameters like the ratios of fatty acids within the plaster. However, the reliability of these ratios for binder identification is severely limited, as demonstrated in this manuscript. Therefore, a more reliable tool for effective differentiation between egg and oil binders based on a combination of diagnostic values, specific markers (cholesterol oxidation products), and PCA is presented in this study. Reference samples of wall paintings with egg and linseed oil binders with six different pigments were subjected to modern artificial ageing methods and subsequently analysed using two GC-MS instruments. A statistically significant difference (at a 95% confidence level) between the egg and oil binders and between the results from two GC-MS instruments was observed. These discrepancies between the results from the two GC-MS instruments are likely attributed to the heterogeneity of the samples with egg and oil binders. This study highlights the complexities in identifying wall painting binders and the need for innovative and revised analytical methods in conservation efforts.

Conservation Science Department Kunsthistorisches Museum Burgring 5 1010 Vienna Austria

Department of Analytical Chemistry Faculty of Science Palacký University 17 Listopadu 12 779 00 Olomouc Czech Republic

See more in PubMed

Sotiropoulou S., Sciutto G., Tenorio A.L., Mazurek J., Bonaduce I., Prati S., Mazzeo R., Schilling M., Colombini M.P. Advanced Analytical Investigation on Degradation Markers in Wall Paintings. Microchem. J. 2018;139:278–294. doi: 10.1016/j.microc.2018.03.007. DOI

Geddes Da Filicaia E., Evershed R.P., Peggie D.A. Review of Recent Advances on the Use of Mass Spectrometry Techniques for the Study of Organic Materials in Painted Artworks. Anal. Chim. Acta. 2023;1246:340575. doi: 10.1016/j.aca.2022.340575. PubMed DOI

Holclajtner-Antunović I., Stojanović-Marić M., Bajuk-Bogdanović D., Žikić R., Uskoković-Marković S. Multi-Analytical Study of Techniques and Palettes of Wall Paintings of the Monastery of Žiča, Serbia. Spectrochim. Acta Part A Mol. Biomol. Spectrosc. 2016;156:78–88. doi: 10.1016/j.saa.2015.11.031. PubMed DOI

Sfarra S., Ibarra-Castanedo C., Tortora M., Arrizza L., Cerichelli G., Nardi I., Maldague X. Diagnostics of Wall Paintings: A Smart and Reliable Approach. J. Cult. Herit. 2016;18:229–241. doi: 10.1016/j.culher.2015.07.011. DOI

Daffara C., Ambrosini D., Pezzati L., Paoletti D. Thermal Quasi-Reflectography: A New Imaging Tool in Art Conservation. Opt. Express. 2012;20:14746. doi: 10.1364/OE.20.014746. PubMed DOI

Stout S., Cosentino A., Scandurra C. Non-Invasive Materials Analysis Using Portable X-ray Fluorescence (XRF) in the Examination of Two Mural Paintings in the Catacombs of San Giovanni, Syracuse. In: Ioannides M., Magnenat-Thalmann N., Fink E., Žarnić R., Yen A.-Y., Quak E., editors. Digital Heritage. Progress in Cultural Heritage: Documentation, Preservation, and Protection. Volume 8740. Springer International Publishing; Cham, Switzerland: 2014. pp. 697–705. Lecture Notes in Computer Science. DOI

Piqué F., Verri G. Organic Materials in Wall Paintings: Project Report. The Getty Conservation Institute; Los Angeles, CA, USA: 2015.

Borg B., Dunn M., Ang A., Villis C. The Application of State-of-the-Art Technologies to Support Artwork Conservation: Literature Review. J. Cult. Herit. 2020;44:239–259. doi: 10.1016/j.culher.2020.02.010. DOI

Mora P., Mora L., Philippot P. La Conservazione Delle Pitture Murali. 2nd ed. Compositori; Bologna, Italy: 2001.

Cennini C. In: Il Libro Dell’arte. 7th ed. Frezzato F., editor. Neri Pozza; Vicenza, Italy: 2012.

Cennini C., Broecke L. Cennino Cennini’s ‘Il Libro Dell’Arte’: A New English Translation and Commentary with Italian Transcription. Archetype; London, UK: 2015.

Mills J.S., White R. The Organic Chemistry of Museum Objects. 2nd ed. Butterworth-Heinemann Ltd.; Oxford, UK: 1994.

Schilling M., Khanjian H. Gas Chromatographic Determination of the Fatty Acid and Glycerol Content of Lipids. I. The Effects of Pigments and Aging on the Composition of Oil Paints; Proceedings of the ICOM Committee for Conservation Preprints, 11th Triennial Meeting; Edinburgh, UK. 1–6 September 1996; pp. 220–227.

Vallance S.L. Critical Review Applications of Chromatography in Art Conservation: Techniques Used for the Analysis and Identification of Proteinaceous and Gum Binding Media. Analyst. 1997;122:75R–81R. doi: 10.1039/a606219i. DOI

Regazzoni L., Cavallo G., Biondelli D., Gilardi J. Microscopic Analysis of Wall Painting Techniques: Laboratory Replicas and Romanesque Case Studies in Southern Switzerland. Stud. Conserv. 2018;63:326–341. doi: 10.1080/00393630.2017.1422891. DOI

Colombini M.P., Modugno F., Fuoco R., Tognazzi A. A GC-MS Study on the Deterioration of Lipidic Paint Binders. Microchem. J. 2002;73:175–185. doi: 10.1016/S0026-265X(02)00062-0. DOI

Chiavari G., Gandini N., Russo P., Fabbri D. Characterisation of Standard Tempera Painting Layers Containing Proteinaceous Binders by Pyrolysis (/Methylation)-Gas Chromatography-Mass Spectrometry. Chromatographia. 1998;47:420–426. doi: 10.1007/BF02466473. DOI

Silva R.H.D. The Problem of The Binding Medium Particularly in Wall Painting. Archaeometry. 1963;6:56–64. doi: 10.1111/j.1475-4754.1963.tb00580.x. DOI

Casadio F., Giangualano I., Piqué F. Organic Materials in Wall Paintings: The Historical and Analytical Literature. Stud. Conserv. 2004;49((Suppl. S1)):63–80. doi: 10.1179/sic.2004.49.Supplement-1.63. DOI

Kouloumpi E., Lawson G., Pavlidis V. The Contribution of Gas Chromatography to the Resynthesis of the Post-Byzantine Artist’s Technique. Anal. Bioanal. Chem. 2007;387:803–812. doi: 10.1007/s00216-006-0802-x. PubMed DOI

Mills J.S. The Gas Chromatographic Examination of Paint Media. Part I. Fatty Acid Composition and Identification of Dried Oil Films. Stud. Conserv. 1966;11:92–107. doi: 10.1179/sic.1966.011. DOI

Laurie A.P. The Painter’s Methods. Dover Publications, Inc.; Mineola, NY, USA: 1988.

Theophilus . On Divers Arts—The Foremost Medieval Treatise on Painting, Glassmaking and Metalwork. Dover Publications, Inc.; New York, NY, USA: 1979.

van den Berg J.D.J. Analytical Chemical Studies on Traditional Linseed Oil Paints. Molart; Amsterdam, The Netherlands: 2002.

White R., Pilc J. Analyses of Paint Media. Natl. Gallery Technol. Bull. 1996;17:91–103.

Bensi P., Danti C., Matteini M., Moles A. Le Pitture Murali: Tecniche, Problemi, Conservazione. Centro Di; Firenze, Italy: 1990. La Pellicola Pittorica Nella Pittura Murale in Italia: Materiali e Tecniche Esecutive Dall’alto Medioevo al XIX Secolo; pp. 73–102.

Hess M. Hess’s Paint Film Defects, Their Causes and Cure. 3rd ed. Chapman and Hall; London, UK: 1979.

Levison H.W. Yellowing and Bleaching of Paint Films. J. Am. Inst. Conserv. 1985;24:69–76. doi: 10.1179/019713685806028123. DOI

Mills J., White R. Organic Mass-Spectrometry of Art Materials: Work in Progress. Natl. Gallery Technol. Bull. 1982;6:3–18.

Manzano E., Rodriguez-Simón L.R., Navas N., Checa-Moreno R., Romero-Gámez M., Capitan-Vallvey L.F. Study of the GC–MS Determination of the Palmitic–Stearic Acid Ratio for the Characterisation of Drying Oil in Painting: La Encarnación by Alonso Cano as a Case Study. Talanta. 2011;84:1148–1154. doi: 10.1016/j.talanta.2011.03.012. PubMed DOI

Schilling M., Khanjian H., Carson D. Fatty Acid and Glycerol Content of Lipids; Effects of Ageing and Solvent Extraction on the Composition of Oil Paints. J. Technol. Archit. Environ. 1997;5:71–78.

Manzano E., Rodríguez-Simón L.R., Navas N., Capitán-Vallvey L.F. Non-Invasive and Spectroscopic Techniques for the Study of Alonso Cano’s Visitation from the Golden Age of Spain. Stud. Conserv. 2021;66:298–312. doi: 10.1080/00393630.2020.1830528. DOI

Colombini M.P., Modugno F., editors. Organic Mass Spectrometry in Art and Archaeology. Wiley; Hoboken, NJ, USA: 2009. Organic Materials in Art and Archaeology; pp. 1–36. DOI

Colombini M.P., Andreotti A., Bonaduce I., Modugno F., Ribechini E. Analytical Strategies for Characterizing Organic Paint Media Using Gas Chromatography/Mass Spectrometry. Acc. Chem. Res. 2010;43:715–727. doi: 10.1021/ar900185f. PubMed DOI

Bonaduce I., Carlyle L., Colombini M.P., Duce C., Ferrari C., Ribechini E., Selleri P., Tiné M.R. New Insights into the Ageing of Linseed Oil Paint Binder: A Qualitative and Quantitative Analytical Study. PLoS ONE. 2012;7:e49333. doi: 10.1371/journal.pone.0049333. PubMed DOI PMC

Schilling M., Carson D., Khanjian H. Gas Chromatographic Determination of the Fatty Acid and Glycerol Content of Lipids. IV. Evaporation of Fatty Acids and the Formation of Ghost Images by Framed Oil Paintings; Proceedings of the ICOM Committee for Conservation Preprints, 12th Triennial Meeting; Lyon, France. 29 August–3 September 1999; pp. 242–247.

Tsakalof A.K., Bairachtari K.A., Chryssoulakis I.D. Pitfalls in Drying Oils Identification in Art Objects by Gas Chromatography. J. Sep. Sci. 2006;29:1642–1646. doi: 10.1002/jssc.200500411. PubMed DOI

Rampazzi L., Cariati F., Tanda G., Colombini M.P. Characterisation of Wall Paintings in the Sos Furrighesos Necropolis (Anela, Italy) J. Cult. Herit. 2002;3:237–240. doi: 10.1016/S1296-2074(02)01180-9. DOI

Tammekivi E., Vahur S., Vilbaste M., Leito I. Quantitative GC–MS Analysis of Artificially Aged Paints with Variable Pigment and Linseed Oil Ratios. Molecules. 2021;26:2218. doi: 10.3390/molecules26082218. PubMed DOI PMC

Pitthard V., Griesser M., Stanek S. Methodology and Application of GC-MS to Study Altered Organic Binding Media from Objects of the Kunsthistorisches Museum, Vienna. Ann. Chim. 2006;96:561–573. doi: 10.1002/adic.200690058. PubMed DOI

Wei S. Ph.D. Thesis. Vienna University of Technology; Vienna, Austria: 2007. A Study of Natural Organic Binding Media Used in Artworks and of Their Ageing Behaviour by GC/FID and GC/MS.

Pitthard V., Finch P., Bayerová T. Direct Chemolysis-Gas Chromatography-Mass Spectrometry for Analysis of Paint Materials. J. Sep. Sci. 2004;27:200–208. doi: 10.1002/jssc.200301617. PubMed DOI

Colombini M.P., Modugno F., Giacomelli M., Francesconi S. Characterisation of Proteinaceous Binders and Drying Oils in Wall Painting Samples by Gas Chromatography–Mass Spectrometry. J. Chromatogr. A. 1999;846:113–124. doi: 10.1016/S0021-9673(99)00344-1. DOI

Schlenk H., Gellerman J.L. Esterification of Fatty Acids with Diazomethane on a Small Scale. Anal. Chem. 1960;32:1412–1414. doi: 10.1021/ac60167a011. DOI

Glastrup J. A Note on the Analysis of the Binding Medium from a Phoenician Shipwreck. Stud. Conserv. 1995;40:65–68. doi: 10.1179/sic.1995.40.1.65. DOI

Blau K., Halket M.J. Handbook of Derivatives for Chromatography. 2nd ed. John Wiley & Sons; Chichester, UK: 1993.

Seppänen-Laakso T., Laakso I., Hiltunen R. Analysis of Fatty Acids by Gas Chromatography, and Its Relevance to Research on Health and Nutrition. Anal. Chim. Acta. 2002;465:39–62. doi: 10.1016/S0003-2670(02)00397-5. DOI

Eder K. Gas Chromatographic Analysis of Fatty Acid Methyl Esters. J. Chromatogr. B Biomed. Sci. App. 1995;671:113–131. doi: 10.1016/0378-4347(95)00142-6. PubMed DOI

Pitthard V., Stanek S., Griesser M., Muxeneder T. Gas Chromatography—Mass Spectrometry of Binding Media from Early 20th Century Paint Samples from Arnold Schönberg’s Palette. Chromatographia. 2005;62:175–182. doi: 10.1365/s10337-005-0595-7. DOI

Challinor J.M. Review: The Development and Applications of Thermally Assisted Hydrolysis and Methylation Reactions. J. Anal. Appl. Pyrolysis. 2001;61:3–34. doi: 10.1016/S0165-2370(01)00146-2. DOI

MACHEREY-NAGEL GmbH & Co., KG [(accessed on 14 February 2024)]. Available online: https://www.mn-net.com/gc-column-nonpolar-low-polar-optima-5-ms-30-m-l-0.25-mm-id-0.25-m-df-726220.30.

Agilent Technologies. [(accessed on 14 February 2024)]. Available online: https://www.agilent.com/en/product/gc-columns/low-bleed-gc-ms-columns/db-5ms-columns.

Izzo F.C. Ph.D. Thesis. Ca’ Foscari University; Venice, Italy: 2011. 20th Century Artist’s Oil Paints: A Chemical-Physical Survey.

Colombini M.P., Modugno F., Menicagli E., Fuoco R., Giacomelli A. GC-MS Characterization of Proteinaceous and Lipid Binders in UV Aged Polychrome Artifacts. Microchem. J. 2000;67:291–300. doi: 10.1016/S0026-265X(00)00075-8. DOI

Degano I., La Nasa J., Ghelardi E., Modugno F., Colombini M.P. Model Study of Modern Oil-Based Paint Media by Triacylglycerol Profiling in Positive and Negative Ionization Modes. Talanta. 2016;161:62–70. doi: 10.1016/j.talanta.2016.08.017. PubMed DOI

Pitthard V., Griesser M., Stanek S., Bayerova T. Study of Complex Organic Binding Media Systems on Artworks Applying GC-MS Analysis: Selected Examples from the Kunsthistorisches Museum, Vienna. Macromol. Symp. 2006;238:37–45. doi: 10.1002/masy.200650606. DOI

Antova G.A., Gerzilov V.T., Petkova Z.Y., Boncheva V.N., Bozhichkova I.N., St Penkov D., Petrov P.B. Comparative Analysis of Nutrient Content and Energy of Eggs from Different Chicken Genotypes. J. Sci. Food Agric. 2019;99:5890–5898. doi: 10.1002/jsfa.9863. PubMed DOI

Andreotti A., Bonaduce I., Colombini M.P., Gautier G., Modugno F., Ribechini E. Combined GC/MS Analytical Procedure for the Characterization of Glycerolipid, Waxy, Resinous, and Proteinaceous Materials in a Unique Paint Microsample. Anal. Chem. 2006;78:4490–4500. doi: 10.1021/ac0519615. PubMed DOI

Serefidou M., Bracci S., Tapete D., Andreotti A., Biondi L., Colombini M.P., Giannini C., Parenti D. Microchemical and Microscopic Characterization of the Pictorial Quality of Egg-Tempera Polyptych, Late 14th Century, Florence, Italy. Microchem. J. 2016;127:187–198. doi: 10.1016/j.microc.2016.03.001. DOI

Mills J.S., White R. The Identification of Paint Media from the Analysis of Their Sterol Composition: A Critical View. Stud. Conserv. 1975;20:176. doi: 10.2307/1505737. DOI

Van Den Brink O.F., Ferreira E.S.B., Van Der Horst J., Boon J.J. A Direct Temperature-Resolved Tandem Mass Spectrometry Study of Cholesterol Oxidation Products in Light-Aged Egg Tempera Paints with Examples from Works of Art. Int. J. Mass. Spectrom. 2009;284:12–21. doi: 10.1016/j.ijms.2008.11.005. DOI

Boon J.J., Peulvé S., Van Den Brink O.F., Duursma M., Rainford D. Molecular Aspects of Mobile and Stationary Phases in Ageing Tempera and Oil Paint Films. In: Bakkenist T., Hoppenbrouwers R., Dubois H., editors. Early Italian Paintings—Techniques and Analysis. Limburg Conservation Institute; Maastricht, The Netherlands: 1997. pp. 35–56.

Buckley S.A., Stott A.W., Evershed R.P. Studies of Organic Residues from Ancient Egyptian Mummies Using High Temperature-Gas Chromatography-Mass Spectrometry and Sequential Thermal Desorption-Gas Chromatography-Mass Spectrometry and Pyrolysis-Gas Chromatography-Mass Spectrometry. Analys. 1999;124:443–452. doi: 10.1039/a809022j. PubMed DOI

Herchi W., Harrabi S., Sebei K., Rochut S., Boukhchina S., Pepe C., Kallel H. Phytosterols Accumulation in the Seeds of Linum usitatissimum L. Plant Physiol. Biochem. 2009;47:880–885. doi: 10.1016/j.plaphy.2009.07.001. PubMed DOI

Rubner-Institut M. Fatty Acid Composition, Tocopherol and Sterol Contents in Linseed (Linum usitatissimum L.) Varieties. Iran. J. Chem. Chem. Eng. 2017;36:147–152.

Colombini M.P., Modugno F., Giacomelli A. Two Procedures for Suppressing Interference from Inorganic Pigments in the Analysis by Gas Chromatography–Mass Spectrometry of Proteinaceous Binders in Paintings. J. Chromatogr. A. 1999;846:101–111. doi: 10.1016/S0021-9673(99)00192-2. DOI

Cennini d’Andrea C., Thompson D.V. The Craftman’s Handbook: The Italian “Il Libro Dell’Arte”. Dover Publications, Inc.; New York, NY, USA: 1954.

Jarque C.M., Bera A.K. Efficient Tests for Normality, Homoscedasticity and Serial Independence of Regression Residuals. Econ. Lett. 1980;6:255–259. doi: 10.1016/0165-1765(80)90024-5. DOI

Jarque C.M., Bera A.K. A Test for Normality of Observations and Regression Residuals. Int. Stat. Rev./Rev. Int. Stat. 1987;55:163. doi: 10.2307/1403192. DOI

Kolmogorov A. Sulla Determinazione Empirica Di Una Legge Di Distribuzione. G. Dell’ist. Ital. Degli Attuari. 1933;4:83–91.

Smirnov N. Sur Les Ecarts de La Courbe de Distribution Empirique. Recl. Mathématique (Mat. Sb.) 1939;6:3–26.

Poulin J. A New Methodology for the Characterisation of Natural Dyes on Museum Objects Using Gas Chromatography–Mass Spectrometry. Stud. Conserv. 2018;63:36–61. doi: 10.1080/00393630.2016.1271097. DOI