3.1. Optical, scanning electron and atomic force microscopy
Figure 2a shows the image of the sample excised from an altered blue area of the painting. It can be seen that the paint layer exhibits a notable darkening in the surface as consequence of the alteration processes that took place on the painting.
The examination of the cross-section of the altered and unaltered paint samples under the optical microscope allowed the recognition of the layered structure and the morphological features. The stratigraphic distribution of the paint samples examined comprises two light-blue paint layers and two reddish ground layers (Fig. 2b). The upper blue paint layer is thinner (at ca 15 µm) than the lower one (at ca 55 µm) and is composed of blue pigment. In the upper blue paint layer can be seen microparticles and dark-blue aggregates with variable sizes (1–5 µm) of blue pigments. In the inner paint layer can be seen heterometric rounded lead white particles distributed heterogeneously throughout the light-blue microcrystalline matrix. Interestingly, the chromatic alteration observed in the painting is observed in the outer 5 µm of the upper blue paint layer in the cross-section (Fig. 2b), suggesting that the alteration process is a surface phenomenon as confirmed in other studies [19, 44]. The lower red ground layer presents a heterogeneous texture in which translucent, white and red-ochre small grains are dispersed in a reddish matrix and the upper red ground layer shows similar characteristics but a thinner texture. Some other morphological features can be seen in the cross-section, such as microcracks across the paint layers, partial loss of the outer paint layer, cavities between the ground and the paint layers, and a disjunction between both ground layers, which highlight the deterioration of this paint sample.
Backscattered electron (BSE) images and analysis performed by energy dispersive X-ray spectrometry (EDX) on this sample revealed in the lower paint layer well-defined light gray particles (highly scattering) where lead is detected, corresponding to lead white (2PbCO3·Pb(OH)2), as can be seen in Fig. 2c. Furthermore, dark-grey (less scattering) amorphous areas and small particles are also identified in the upper paint layer in which iron is detected in significant proportion, along with potassium (Fig. 2d). The abundance of iron in most part of the upper layer together with potassium suggest the presence of Prussian blue (FeIII4[FeII(CN)6]3 or KFeIII[FeII(CN)6].xH2O, depending on the preparation procedure). On the other hand, the presence of iron together with aluminium, sulphur and potassium appear to be characteristic of the original method of Prussian blue preparation in which dried blood or another animal matter and alum (aluminium potassium sulphate) as extender are ingredients, following the procedure reported by Woodward [45]. However, in some of these dark grey amorphous areas the absence of iron is evident (results not shown) and this feature could suggest the coexistence of an organic pigment such as indigo, precipitated in alum for shading, and/or a deterioration of the Prussian blue pigment due to a lixiviation of the iron ions/solubilization of the Prussian blue. Finally, ground layers show a typical profile of a red earth pigment due to the identification of aluminum, silicon, magnesium, potassium, calcium and iron (results not shown), but the abundance of lead indicates the possible addition of red lead or the diffusion of lead-compounds (such as lead soaps) from the paint upper layers.
Figure 3 shows the AFM topographic images (10 × 10) µm obtained in samples excised from altered (Fig. 3a) and unaltered (Fig. 3b) areas of the painting. It can be seen that the micromorphology in both cases is similar with pigment grains protruding the surface resulting in Δhigh in the range of 1 µm. It results more interesting the comparison between the elastic modulus obtained for paint samples excised from unaltered and altered areas of the painting which are summarized in Table 1. It is firstly worth of mention that the EM values obtained in this research were in the same order as those for the 19th -century oil paintings reported in other studies that focused on changes in nanomechanical properties while aging oil paintings. [46]. The higher values obtained for the EM in the altered sample were associated with increased stiffness due to the alteration processes taking place in the painting. As described in sections thereafter, these alteration processes undergone by the painting have resulted in the hydrolysis of the drying oil and the concomitant depolymerisation and loss of the cross-linking of the oil network. Consequently, the paint film has undergone an increase of stiffness.
Table 1
Values of the elastic modulus obtained by means of NI-AFM in microsamples of unaltered and altered paint film excised from the “San Francesc de Paula” painting.
Elastic modulus | Unaltered blue paint (GPa) | Altered blue paint (GPa) |
Mean Value | 6.9 | 10.2 |
Minimum value | 6.0 | 6.3 |
Maximum value | 9.4 | 20.4 |
3.2. FTIR spectroscopy
The results of the FTIR analysis of the altered paint sample (Fig. 4) evidenced the predominance of absorption bands ascribed to lead carbonate in the basic or hidrocerussite form (2PbCO3.Pb(OH)2) (absorption bands at 3536, 1393, 1045 and 679 cm−1). On the other hand, the absorption bands appearing in the region between 3000 − 2900 cm−1 are related to stretching vibrations of methyl and methylene groups from fatty acid chains, demonstrating the presence of an organic material of lipidic type as binding media. This organic material exhibits an advanced state of degradation, as confirmed by the identification of a carboxylic acid band related to the free fatty acids at about 1703 cm−1, as well as by the spectral region between 1650 − 1500 cm−1, with characteristic bands attributed to metal carboxylates, mainly lead carboxylates (band at 1506 and shoulder at 1537 cm−1) formed between the free fatty acids and the lead ions of the pigment, and oxalates of calcium (weddellite and whewellite at 1643 and 1620 cm−1) and lead (1651 cm−1) [14]. In this sense, it should be pointed out that calcium oxalates are more abundant in the altered paint sample than in the unaltered one (results not shown) and these compounds can contribute to the chromatic alterations showed in the painting. Other inorganic compounds were also identified in the infrared spectrum such as calcite (CaCO3: 1400, 871 cm−1), siliceous minerals ascribed to earth pigments (1166, 1026, 797 and 767 cm−1) and alum (1094 cm−1).
A low intensity absorption band with a maximum at 2089 cm−1 related to the stretching vibration of CN group confirms the presence of Prussian blue in accordance with the results of SEM/EDX. The broadening of this band and the shoulder at 2050 cm−1 suggest the coexistence of different species with variated oxidation states formed through redox reactions of this pigment [47]. However, the considerable overlap between the absorption bands of the organic and inorganic compounds present in the pain samples due to the complexity of this matrix make difficult the confirmation of the coexistence of other pigments such as indigo.
Figure 5 shows the IR absorption spectra of the reconstructed pain specimens made with Prussian blue (PO) (Fig. 5a), indigo (IO) (Fig. 5b) and the mixture 1:1 indigo-Prussian (IPO) (Fig. 5c). The IR spectrum of Prussian blue + oil paint film is dominated by strong band at 2082 cm−1 ascribed to the stretching vibration of CN group. Indigo + oil paint film exhibits abundant IR bands in the fingerprint region dominated by the band at 1069 cm−1 and moderate bands at 1625 and 1610 cm−1 ascribed to the stretching vibration of C = O group in indigoid molecules present in natural indigo. Linseed oil is recognized in both IR spectra by sharp bands at 2924 and 2853 cm−1 associated with the stretching vibrations of methyl and methylene groups in triacylglycerol molecules and by the band at 1738 cm−1 ascribed to the carbonyl bond in the ester groups of triacylglycerol molecules. The IR spectrum of the indigo-Prussian blue + oil reconstructed paint film is a combination of the IR spectra of both pigments plus the drying oil. This last spectrum is dominated by the Prussian blue band of CN groups at 2082 cm−1. Moderate bands of carbonyl stretch at 1738 cm−1 and C-O stretch vibrations at 1240, 1158 and 1097 cm−1 ascribed to the linseed oil are also present. Interestingly a shoulder at 1705 cm−1, ascribed to stretching vibrations of free fatty acids released by the hydrolysis reaction undergone by the triacylglycerol molecules of the linseed oil is exclusively observed in reconstructed paint films containing Prussian blue.
Figure 6a shows the IR spectra of the indigo-Prussian blue + oil reconstructed paint film (IPO) (IR spectrum a) after ageing and the reconstructed paint films (IPO-N, IPO-UVA) after natural (IR spectrum b) and UV solar light (IR spectrum c). It can be recognized changes in spectral region I (1800 − 1600 cm−1) and spectral region II (1100 − 1000 cm−1). These changes in IR bands can be seen in detail in Fig. 6b. Interestingly a new band at 1610 cm−1 ascribed to carbonyl band in indigoid molecules as well as a new band at 1073 cm−1 ascribed to stretching vibrations of OH groups occurs in both natural and UV solar light aged paint films.
3.3 Gas chromatography-mass spectrometry
With the purpose of completely characterizing the organic material used as binding media in the painting, and thus, complementing the results obtained by ATR-FTIR, representative fragments of the altered and unaltered paint samples were analyzed by means of GC-MS. In Fig. 7 is shown the chromatogram obtained for the altered paint sample that revealed the presence of monocarboxylic even-numbered saturated fatty acids (peaks 4–5) containing 16 and 18 carbon atoms, with stearic acid (18:0) as the most abundant, and short-chain dicarboxylic acids with 8, 9 and 10 carbon atoms (peaks 1–3) in a significant proportion formed due to oxidation process undergone by the unsaturated fatty acids (oleic, linoleic and linolenic). A similar chromatographic profile was obtained for the paint sample from the unaltered area (results not shown).
The occurrence of these compounds confirms the presence of an organic material of lipid type as binder. On the other hand, taken into account these results, the P/S ratio (chromatographic peak area of palmitic acid versus chromatographic peak area of stearic acid) obtained is 0.6–0.7, which is not adjusted for the expected values for linseed, walnut, poppy or mixtures of them, commonly used in oil paintings [48]. This deviation indicates significant variations in the content of fatty acids constituent of the drying oil as a consequence of the high degree of degradation of the binding media, which has undergone an extensive hydrolysis process and the subsequent formation of metal soaps between the free-fatty acids of the oil and the metal ions of the pigments, as well as oxalates, as shown in the results obtained by the ATR-FTIR analysis.
3.4. Voltammetric pattern
Figure 8 shows the square wave voltammograms of microparticulate deposits of a) indigo and b) isatin abrasively transferred onto the surface of graphite electrodes in contact with 0.25 M HAc/NaAc aqueous buffer at pH 4.75. These are accompanied by the voltammograms corresponding to c) unaltered and d) altered layers of the Sant Francesc de Paula paint. Upon scanning the potential in the negative direction, indigo (Fig. 8a) displays two well-defined peaks at −0.30 (I) and 0.45 V vs. Ag/AgCl (II) corresponding, respectively, to the reduction of indigo (IND) to leucoindigo (LEU) and the oxidation of indigo to dehydroindigo (DHI). These two proton-assisted solid-state processes are summarized in Scheme 1 and can be represented as [36–38]:
{IND} solid + 2H+ aq + 2e− → {LEU} solid (1)
{IND} solid → {DHI} solid + 2H+ aq + 2e− (2)
In turn, isatin produces a unique, intense cathodic peak at ca. −0.70 V (III) (Fig. 8b), corresponding to the proton-assisted reduction to hydroxylated derivatives (see scheme 1) [39, 49].
The unaltered blue submicrosample (Fig. 8c) yields the indigo-localized peaks I and II accompanied by broad peaks at −0.15 (IV) and 0.80 V (V), which can be unambiguously attributed to Prussian blue centered processes on the basis of abundant literature on the electrochemistry of this compound [33–35] and our blank experiments in synthetic specimens (vide infra). These processes correspond, respectively, to the reduction of Fe(II) centers and the oxidation of Fe(III) ones coupled to the entrance/issue of electrolyte charge-balancing cations [33–35]. Signals attributable to lead white were recorded only in positive-going voltammograms appearing as weak anodic signals at ca. −0.50 V, in agreement with prior studies [50, 51]. The voltammetric peaks IV and V can be represented as [33–35]:
KFeIII[FeII(CN)6] solid + K+ aq + e− → K2FeII[FeII(CN)6] solid (3)
KFeIII[FeII(CN)6] solid → FeIII[FeIII(CN)6] solid + K+ aq + e− (4)
Interestingly, the voltammograms of submicrosamples extracted from the altered regions of the paint show intense signals III and IV while the indigo signals and the Prussian blue signal V vanish (Fig. 8d). These features suggest that there is an oxidation of indigo to isatin and Prussian blue to Berlin green responsible for the chromatic change of the paint.
3.5. Degradation pathways
In order to study the possible mechanisms of degradation, specimens of indigo, Prussian blue, 50%wt mixture of these pigments and reconstructed model paint films containing either indigo, Prussian blue, 50%wt mixture of these pigments dispersed into linseed oil (see experimental section), were submitted to two aging protocols, natural ageing for 2 years and accelerated ageing with UVA (UV solar light) for 200 h. The most relevant voltammetric features are summarized in Figs. 9 and 10. The former superimposes the negative-going potential scan voltammograms of indigo, Prussian blue and o 50% wt mixture of both components, clearly showing the indigo-centered signals I and II and the Prussian blue centered peaks IV and V.
In Fig. 10a, the electrochemical response of the parent pigment blue mixture is superimposed to those of the mixture submitted to natural and UV aging, while Fig. 10b compares the voltammograms of a paint film specimen and those submitted to natural and UV solar light aging. The voltammetric features of pigments and paint specimens after natural and UV solar light aging were essentially identical. The most relevant features can be summarized as:
a) In the pure pigments, as well as in the pigment mixtures, natural aging determines a decrease of the indigo signal II without concomitant decrease of the signal I, a light increase in the Prussian blue signal IV without variation of the signal V, all these features being accompanied by the appearance of a new voltammetric peak at 1.05 V (VI). Upon natural and UV solar light aging, the decrease of the indigo signal becomes more pronounced while the peak VI becomes clearly enhanced. This last signal can tentatively be attributed to a species resulting from the hydroxyl addition to the indigo molecule (see Scheme 1) whose electrochemical oxidation should occur at high potentials.
b) The paint oil specimens show the Prussian blue signal V clearly depleted relative to the indigo signals. Natural aging determines the same features described for the pure pigments, but here the Prussian blue signals become enhanced. Under UV aging, the peak VI is considerably enhanced whereas both indigo peaks I and II are depleted.
c) In all indigo-containing specimens, the isatin signature (peak III) appears upon aging, being particularly enhanced when UV aging was employed.
These voltammetric data are consistent with infrared features. As can be seen in Fig. 5b, appearance of a new stretch band of C = O groups in indigoid molecules at 1605 cm−1 on UV and UV solar light ageing suggests diversification of indigoid molecules on ageing. Similarly, the new indigo IR band at 1073 cm−1, which has been tentatively ascribed to OH groups, also suggests a diversification of indigoid molecules as result of the alteration processes that have taken place. Appearance of the carbonyl signal at 1705 cm−1 only in reconstructed paint films that contain Prussian blue (PO and IPO), suggests that the hydrolysis process undergone by the linseed oil is promoted by the Prussian blue pigment.
These features can be interpreted on the basis of studies on the degradation of indigo and Prussian blue pigments. In regard to indigo, its degradation is in principle promoted by oxidants [41], the degradation by light being, however, particularly intense [42]. According to Iuga et al. [43], there are two (main) indigo degradation pathways catalyzed by hydroxyl (•OH) and hydroperoxyl (•OOH) radicals. In turn, the hydroxyl-catalyzed degradation can lead to dehydroindigo via intermediate formation of an indigo radical anion after proton abstraction, or via OH addition to the central C = = C indigo bond. Ultimately, indigo photodegradation yields isatin, isatoic anhydride and anthranilic acid [41–43, 52] whereas indigo degradation by ozone yields isatin and isatoic anhydride [53].
In regard to Prussian blue, Samain et al. [20] have reported that it can be partially oxidized to Berlin green (KFeIII[FeII(CN)6])x(FeIII[FeIII(CN)6])1−x. The degradation process appears to be facilitated in the presence of linseed oil binder and lead white, leading to the reduction of Prussian blue to Berlin white, K2FeII(FeII(CN)6, at the exposed paint surface and an oxidation to Berlin green in the bulk of the paint layer [20]. Then, the combination of isatin and other indigo degradation products and Berlin green can be responsible for the observed chromatic alteration in the Sant Francesc de Paula painting.
On the other hand, the polymerization of drying oils produces free radicals (alcoxyle, •OR, peroxyle, •OOR) derived from hydroperoxides [21], able to promote the above oxidation reactions. Accordingly, the formation of isatin, and hence, the orange-brownish hue acquired by the degraded zones of the Sant Frances de Paula paint, can be attributed to the oxidation of indigo promoted by free radicals generated in the oil binding. As previously noted, this situation is mainly reproduced in paint specimens prepared from indigo without Prussian blue. To interpret the minor presence or even the absence of isatin in aged specimens containing both Prussian blue and indigo can be rationalized on considering the peculiar characteristics of the interaction of Prussian blue with reactive oxygen species (ROS). This compound acts as catalyst for H2O2 reduction under electrochemical conditions [54, 55] where it appears to be partially dissolved releasing Fe2+ (aq) and ferrocyanide associated to local production of HO−, so that Fe2+ (aq) ions initiate the Fenton reaction with H2O2 generating hydroxyl radicals [56]. On the contrary, it has been reported that under ‘chemical’ conditions, Prussian blue nanoparticles act as radical scavengers, in particular abstracting •OH radicals mimicking the activity of peroxidases [57, 58].
On the basis of the foregoing set of considerations, the different degradation pathways observed in synthetic paint film specimens and localized areas of the Sant Francesc de Paula paint can be interpreted taking into account that the grain size and the nature of the local binding environment can affect significantly the stability of the pigments [52, 59]. Under ‘ordinary’ conditions of degradation, as is the case of the prepared paint film specimens, Prussian blue which exerts an effect of moderator, acting as radical scavenger [57, 58], being slowly oxidized to Berlin green and leading indigo degradation to the formation of adducts with the radicals, as schematically depicted in Scheme 1.
In the case of the Sant Francesc de Paula paint, since the canvas was rolled up and re-mounted onto a wooden stretcher and subsequently re-varnished [32], the produced fissures provided opportunity for a reinforced action of light and humidity on the pigment layers. Here, Prussian blue acts as an initiator of the Fenton reaction [56] rather than a radical scavenger and promotes the oxidation of indigo to isatin. This process formally results from the reaction of indigo with O2, or, possibly, with H2O2 formed, in the absence of radical scavenging, by condensation of hydroxyl radicals. Scheme 2 shows an idealized scheme of the possible processes involved in pigment degradation.