Revealing new insights into black stains on the passepartout of Leonardo da Vinci’s Codex Atlanticus

The Codex Atlanticus is one of the largest collections of Leonardo da Vinci's drawings and writings and is currently preserved at the Biblioteca Ambrosiana in Milan. Following restoration, carried out at Grottaferrata between 1962 and 1972, the manuscript consists of 1119 original fragments or folii, each framed with a modern passepartout added by the restorers to allow consultation of both sides of the documents. In 2006, greyish-black stains were discovered on approximately 210 of the passepartouts, around 1-2 cm from the folii, causing concern among curators and conservators. Microbiological research has ruled out any kind of microbial attack on the Codex as the origin of the dark stains.  

With the help of the museum curator at the Biblioteca Ambrosiana, a group of researchers from Politecnico di Milano (Italy) and the ESRF have conducted an extensive analysis of Folio 843 of the Codex to better understand the blackening process. Non-invasive imaging measurements of the folio, micro-imaging and synchrotron spectroscopy investigations of passepartout fragments at different magnifications and spectral energy ranges were exploited to examine the glued modern paper and the composition of the black stains.

The original paper margins could be observed thanks to photoluminescence hyperspectral lifetime imaging (PL-HIS), which demonstrated that the black stains were not made of fluorescent materials. Micro-attenuated total reflection Fourier transform infrared (μATR-FTIR) imaging of fragments from the passepartout revealed the presence of a mixture of starch and PVAc glues, localised only in the stained areas close to the margin of the folio. Observations using field-emission scanning electron microscopy revealed that the black stains were concentrated inside spaces between cellulose fibres, where an aggregation of inorganic, roundish particles made of Hg and S were found to be present.

A variety of synchrotron radiation-based X-ray investigations were carried out to characterise the small particles that made up the dark stains. Synchrotron-based X-ray fluorescence (XRF) mapping and X-ray absorption near-edge structure (XANES) data were acquired at beamline ID21. Macro and µXRF maps supported earlier findings from X-ray spectroscopy analyses, inferring the presence of Hg, S, Ca, Al, Cl, Si and K. Mercury was detected in the darkened region, while it was not detected in the region without alteration. A similar distribution was found for sulfur, although it was also detected in the white regions. µXRF mapping of a stain showed a co-localisation of Hg and S that follows the shape of the dark stain almost perfectly (Figure 1a,b,d).

Click image to enlarge

Fig. 1: a) Visible light image of the black stain region region where µXRF mapping was performed [step size (h × v), 4 × 4 μm; map size (h × v), 800 × 800 μm; exp. time, 100 ms/pixel; energy: 2.825 keV]. b) RGB composite of µXRF maps of Hg (red), Cl (green) and S (blue). The magenta colour indicates the region where mercury and sulfur signals overlap, following the shape of the black stain. c) Macro-XANES spectra recorded at the S K-edge. Peak at 2.4737 keV corresponds to the sulfides (S^-II), peak at 2.4777 keV to the sulfites (S^IV) and peak at 2.4839 keV to the sulfates (S^VI). d) Detail of RGB composite of µXRF maps of Hg (red), Cl (green) and S (blue) [step size (h × v), 4 × 4 μm]. e) Selected area of interest for hyperspectral 2D µXRF maps at S K-edge. The RGB composite image shows the distribution of sulfides (S^-II) and sulfates (S^VI) species.

Macro-XANES spectra acquired at the S K-edge (Figure 1c) exhibited a remarkable difference between darkened and non-altered areas, both in signal intensity (high in dark, low in white region) and sulfur oxidation states (mainly sulfates (S^VI) and sulfites (S^IV) in white, also sulfides (S^-II) in the dark region), suggesting a predominant formation of sulfides associated with the blackening (Figure 1e).

Two types of X-ray diffraction (XRD) measurements were performed within the framework of the ESRF’s “Historical Materials block allocation group (BAG)” beamtime access (www.esrf.fr/BAG/HG172). High angular resolution XRD (HR-XRD) measurements were collected at beamline ID22 on samples from the dark and white regions, and μXRD maps were acquired at beamline ID13. Thanks to these observations, the spatial distribution of the black stains and metacinnabar (β-HgS) peak heights could be correlated (Figure 2).

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Fig. 2: a) HR-XRD spectra patterns of the white (grey) and dark (black) regions. Squares and diamonds highlight the peaks related to cellulose and β-HgS respectively. b) Visible light photograph of the area of the dark sample where µXRD mapping was performed [step size (h × v), 3 × 3 μm; map size (h × v), 1.2 × 1.2 mm; exp. time, 10 ms/pixel; energy: 13 keV]. c) µXRD images calculated by integrating XRD intensity over regions of interest of XRD pattern, shown as green and purple rectangles in (a) for cellulose and metacinnabar respectively.

The research highlighted evidence of the spontaneous formation of β-HgS particles in conservation conditions (i.e., controlled environmental conditions including absence of light and low humidity). A first hypothesis about Hg provenance is based on the possible use of anti-vegetative mercury salts (HgCl₂) in the gluing mixture. However, it is important to stress that this compound was not found throughout the current research.  Additionally, it remains to determine if paper hydrolysis and the degrading processes of the glue mixture revealed by ATR-FTIR imaging may have contributed to the blackening phenomenon. Further studies are required to propose a robust mechanism of the formation of β-HgS and to verify if the exposure of HgCl₂ to different sulfur sources, in mild environmental conditions, could lead to the formation of metacinnabar black particles.

Principal publication and authors
Imaging and micro-invasive analyses of black stains on the passepartout of Codex Atlanticus Folio 843 by Leonardo da Vinci, N. Guarnieri (a), M. Ghirardello (b), S. Goidanich (a), D. Comelli (b), D. Dellasega (c), M. Cotte (d,e), E. Fontana (f), L. Toniolo (a), Sci. Rep. 13, 4902 (2023); https://doi.org/10.1038/s41598-023-31129-2
(a) Department of Chemistry, Materials and Chemical Engineering, Politecnico di Milano, Milan (Italy)
(b) Department of Physics, Politecnico di Milano, Milan (Italy)
(c) Department of Energy, Politecnico di Milano, Milan (Italy)
(d) ESRF
(e) Laboratoire d’Archéologie Moléculaire et Structural (LAMS) CNRS UMR 8220, Sorbonne Université, Paris (France)
(f) Veneranda Biblioteca Ambrosiana, Milan (Italy)