## 1 Introduction
The initial attempts to describe the long-term degradation of fibroin due to high temperatures can be traced back to 1989. During a presentation for the American Chemical Society, [PERSON] demonstrated that a temperature of 150 \({}^{\circ}\)C was the most suitable for fibroin degradation [6]. It is important to note that his experiments were preliminary and focused on various textile and paper materials. Since then, infrared spectroscopic research on the long-term high-temperature degradation of fibroin has primarily focused on investigating the vibrations of C=O and N-H in the spectra's Amide I and II regions [7; 8; 9]. It has been shown that this can mainly assess the oxidation of fibroin [1]. In our previous work, we have proposed to look in the \(\delta_{\text{C-H}}\) region of FTIR spectra and have put forward the hydrolysis estimator as integral of 1318 cm\({}^{-1}\) band, which corresponds to deformation vibrations of groups in a free amino acid, or symmetric bending found in free dicarboxylic amino acids (COO-NH- \(\ldots\)COOH) to band integral of CH\({}_{3}\) bending vibration band located at 1442 cm\({}^{-1}\)[1; 10; 11; 12]. However, the most extensively studied estimator in the literature using infrared techniques is the crystallinity estimator (often confirmed by XRD analysis, as XRD is one of the primary methods used to evaluate the degree of crystallinity in silk samples [13]. In FTIR, the ratio of two Amide I band intensities at 1615 and 1655 cm\({}^{-1}\) (reflecting the relative proportion of the polymer in an organized \(\beta\)-sheet, \(\alpha\)-helix, or random coil arrangement) and the ratio of Amide III bands at 1264 and 1230 cm\({}^{-1}\) can be utilized to calculate the crystallinity index based on similar principles [5; 8; 14; 15; 16; 17; 18; 19]. Alternatively, the growth of the Amide I sub-band centered at 1699 cm\({}^{-1}\) can be monitored [20; 21]. Literature reports indicate that a decrease in crystallinity is often associated with a decline in handling properties, making crystallinity measurement valuable for conservators and museum professionals worldwide [8; 22]. Various attempts have been made to measure the degree of crystallinity in historical silk samples using FTIR and XRD techniques [6; 19; 23; 24; 25; 26; 27]. [PERSON] et al., analyzed XRD patterns and FTIR spectra, revealing fluctuations in crystallinity in ancient samples [9; 28]. More detailed findings from [PERSON] et al., indicate the continued presence of crystalline domains in aged samples, although they exhibit almost complete disorientation in historical samples [29]. In wide-angle X-ray scattering (WAXS) experiments conducted by [PERSON] et al., and ATR-FTIR studies by [PERSON] et al., the loss of the amorphous phase has been identified as a cause of reduced mechanical strength of silk fibers [23; 30]. However, [PERSON] and [PERSON] reported difficulties obtaining XRD reflections from ancient samples [17; 19]. It is essential to acknowledge that the estimation of crystallinity from FTIR spectra can sometimes be distorted by vibrations, such as those from oxidation products that occur during the degradation process, leading to changes in the shape of the Amide bonds and complicating calculations of crystallinity changes. Our previous work was dedicated to studying the degradation process and the change in estimators during thermal degradation. We validated the estimators derived from FTIR measurements using XRD, size exclusion chromatography (SEC), and ultraviolet-visible spectroscopy (UV-Vis). We specifically investigated the oxidation estimator (validated with UV-Vis spectra [1] and color change [31]), the hydrolysis estimator (confirmed with SEC analysis [1]), and three crystallinity estimators. However, only one of the crystallinity estimators showed a similar change trend to the XRD calculations [9]. Consequently, we identified three promising degradation markers, which we used to assess the condition of 10 historical samples dating from the 16 th to 19 th century [32].