*3.4. Infrared Markers of Weld Lake Pigments*

Although infrared spectroscopy has rarely been used to characterize flavonoids, several studies have proven its effectiveness in studying flavonoids-metal complexes [12,13]. Machado et al. [32] did DFT calculations of IR and Raman spectroscopies of hydroxyflavones, and their assignments are summarized in Table 4.

As mentioned above, flavonoids-metal complexes are observed in the infrared spectra of *WL3* and *WL4*. Figure 3 compares the two lake pigments and references of luteolin and luteolin 7-*O*-glucoside and shows the similarity between the infrared spectra, namely between *WL4* and luteolin 7-*O*-glucoside. These data are corroborated by the HPLC-DAD analysis, since this recipe has 53.31% of luteolin 7-*O*-glucoside, when compared with the 30.38% of the recipe *WL3*. The presence of a glucoside in position 7-OH shifted the vibrational frequency from 1186 to 1179 cm−1. The fact that both lakes show lower frequencies, at 1178 and 1173 cm−1, corroborates with the HPLC data. The reference of luteolin 3 ,7-di-*O*-glucoside will provide more insight into the effect of glucosides in the infrared spectra, and further studies are underway.

Regarding the presence of an organometallic complex, the stretching vibration of C=O of luteolin at 1666 cm−<sup>1</sup> is shifted to 1632 cm−1. According to Dong, this shift is characteristic of the existence of a complex [12]. It is the co-ordination of carbonyl oxygen with metal ion bonded to 5-OH group of A ring and 4-CO carbonyl group of C ring [12,33,34]. This is also visible in the stretching at 1613 cm<sup>−</sup>1. Moreover, the OH bending of C<sup>5</sup> shifts from 1509 cm−<sup>1</sup> to 1484-9 cm−1, probably also due to the metal coordination. Another possible indication of metal complexation in positions OH (C5) and CO (C4) is the decrease from 1096 cm−<sup>1</sup> to 1077-68 cm−<sup>1</sup> from the stretching of C3-C4. In fact, the extraction solution of *WL3* had pH ~9.4, the optimum conditions for the metal chelation in the OH at C<sup>5</sup> and the carbonyl at C4. The analysis of luteolin, both as aglycone and with glucosides, complexed with Al3+ is ongoing.

**Figure 3.** Infrared spectra of weld lake pigments *Yellow Lake. Cool tint.* (*WL3*) and *Weld yellow* (*WL4*), and references of Lut and Lut-7-*O*-glu.




**Table 4.** *Cont.*

Annotations: ν—stretching, φ—aromatic ring normal vibrations, δ—in-plane deformation, γ—out-of-plane deformation, Γ—out-of-plane deformation of skeleton atoms, Δ—in-plane deformation of skeleton atoms, ip—in-plane, op—out-of-plane, *s*—symmetric mode.

#### **4. Conclusions**

The W&N 19th century Archive Database has proven, once more, to be an exceptional source of information on 19th-century artists' materials and their commercial preparation, enabling the first study of five W&N manufacturing processes for yellow lake pigments from weld.

This investigation showed that W&N 19th-century methods for preparing weld lake pigments involved extracting the dye in neutral-basic media by adding carbonate compounds (KHCO3 and K2CO3) and complexation of flavonoid compounds was always achieved by the addition of Al3+. Five bright yellow lake pigments were obtained and characterized by a multi-analytical approach. Their chromatographic profiles display as main yellows, luteolin 7-*O*-glucoside (Lut-7-*O*-glu) or both Lut-7-*O*-glu plus luteolin 3 ,7-*O*glucoside (Lut-3',7-*O*-glu). In two of the processes, the presence of gypsum (CaSO4·2H2O) was detected by FTIR, being formed as a by-product of the reaction of alum and CaCO3. This work also offers the first identification of weld lake pigments' characteristic infrared bands: the stretching vibration of C=O at 1632 cm−1, the OH of C5 bending at c. 1484-9 cm<sup>−</sup>1, the stretching of C3-C4 at 1077-68 cm−1, all clear indications of metal complexation in positions OH (C5) and CO (C4) of flavonoid compounds.

The five recipes result in two types of lake pigments: yellows in which a filler, such as gypsum, is present (*WL1* and *WL2*) and yellows in which the lake pigment was found in an aluminate matrix (*WL3* as well as *WL4* and *WL5*). In the first type, the paints' mechanical performance is controlled by the filler [35], and the pigments are more opaque when applied as oil paints. On the other hand, the second type allows the preparation of translucent paints that can be applied as glazes.

The yields obtained experimentally were very similar or better than those of W&N, excluding recipes *WL4* and *WL5*, which were considered experiments. In the future, we intend to investigate variants of the processes and explore the full precipitation of the coloring matter as described by W&N in *WL3*. Moreover, since infrared spectroscopy revealed a powerful technique for the characterization of flavonoids-metal chelation, further work is ongoing with the analysis of other luteolin and apigenin "type-chromophores" complexed with Al3+.

The pigment reconstructions will be fundamental to advancing on degradation studies and supporting analytical methodologies useful for identifying weld lake pigments in artworks, contributing to ensuring effective conservation and authentication procedures.

**Supplementary Materials:** The following are available online at https://www.mdpi.com/2571-940 8/4/1/26/s1.

**Author Contributions:** Conceptualization, P.N. and V.O.; methodology, P.N. and V.O.; synthesis, M.V.; investigation, M.V., P.N., V.O., A.C., M.C.O.; writing—original draft preparation, P.N. and V.O.; writing—review and editing, P.N., V.O., A.C., M.J.M.; funding acquisition, M.J.M. All authors have read and agreed to the published version of the manuscript.

**Funding:** This work was supported by the Associate Laboratory for Green Chemistry-LAQV, which is financed by national funds from FCT/MCTES (UIDB/50006/2020 and UIDP/50006/2020). FCT/MCTES also funded the project "Polyphenols in Art: chemistry and biology hand in hand with conservation of cultural heritage" (PTDC/QUI-OUT/29925/2017), in which this work was developed.

**Institutional Review Board Statement:** Not applicable.

**Data Availability Statement:** Not applicable.

**Conflicts of Interest:** The authors declare no conflict of interest.

### **References**

