Two Byzantine VI century manuscripts known as Vienna Dioskurides and Vienna Genesis, held in the Austrian National Library at Vienna, were analysed with in situ non-invasive techniques. Raman spectroscopy, UV-Vis diffuse reflectance spectrophotometry with optic fibres and X-ray fluorescence spectrometry were used to characterise the palette of these early Middle Ages manuscripts. The analytical study was performed to have a better knowledge on the colourants used by ancient miniature painters, a subject known more on the basis of traditional sources (i.e. medieval treatises) than of analytical evidences. Indeed these illuminated manuscripts are, to the authors' knowledge, among the oldest ever being analysed, so that the colourants found in them can be considered among the oldest evidences of their use. The main feature of Vienna Dioskurides and Vienna Genesis palettes is their richness, exemplified by the simultaneous presence of gold and ultramarine blue; in Vienna Dioskurides cinnabar is also present. Information regarding ultramarine blue is surprising, being the analytical evidence of the use of this precious pigment at least three centuries before its use in Western manuscripts, a feature justified by the fact that the Byzantine Empire was the dominant culture in early Middle Ages in the Mediterranean World. Other colourants include azurite and indigo, red lead, orpiment, red and yellow ochres, while a mixture of blue and yellow colourants, known as vergaut, was used to render green hues. Organic colourants were also used, such as madder and Tyrian purple, the latter employed to dye the parchment of Vienna Genesis.
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