Vibrational Spectroscopy 73 (2014) 79–89 Contents lists available at ScienceDirect Vibrational Spectroscopy jou r n al hom ep age: www.elsevier.com/locate/vibspec Water dependent structural changes of silk from Bombyx mori gland to fibre as evidenced by Raman and IR spectroscopies Aline Percot a,∗ , Philippe Colomban b,∗ , Céline Paris b , Hung Manh Dinh a,1 , Marine Wojcieszak a , Bernard Mauchamp c a Sorbonne Universités, UPMC Univ. Paris 06, UMR 8233, MONARIS, F-75005 Paris, France b CNRS, UMR 8233, MONARIS, F-75005 Paris, France c Biologie Fonctionnelle, Insectes et Interaction (bf2i), INRA/INSA, 69621 Villeurbanne Cedex, France a r t i c l e i n f o Article history: Received 21 February 2014 Received in revised form 6 May 2014 Accepted 7 May 2014 Available online 27 May 2014 Keywords: Silk Gland Fibre Bombyx mori Conformation Water Raman ATR-FTIR a b s t r a c t DSC, attenuated total reflexion infrared (ATR-FTIR), and Raman microspectroscopy were used for the first time in a close to in vivo environment to study ready-to-spin Bombyx mori silkworms. The aim was to understand the change of the fibroin backbone organisation from the gland to the fibre. Raman shifts of the Amide I components reveal a strong change of organisation in the middle part of the hydrated gland, as anticipated previously measured modifications of salts concentrations and pH. Series of bands charac- teristics of the fully hydrated silk disappear, as observed for spider silk, despite the different aminoacid sequence. Confirmation is obtained from IR spectra taking into account the superimposed water com- ponent. The change of the silk–water interaction in the central part of the gland, from a hydrophobic to hydrophilic behaviour, is related to the water content decrease along the gland. pH sensitive carboxylate side chains markers confirm the modification. Fibroin organisation was also studied in the dried gland and in the spun fibre. The fibre extrusion by orients the fibroin chains along the fibre axis, with intercalated water molecules, leading to a material with specific mechanical properties, compared to the amorphous dried gland. © 2014 Published by Elsevier B.V. 1. Introduction Domesticated Bombyx mori (B.m.) silkworm fibre has been stud- ied for many years for large applications in textile and for some extent in biomedical field [1–4] because of its remarkable physical and chemical properties. Silk B.m. fibre exhibits high mechanical strength, strain and work of fracture as well as smooth surfaces and high gloss. From a chemical point of view, silk fibre is a polyamide, an insoluble protein, with side chain grafts corre- sponding to hydrophobic amino acid residues: typically alanine (A: 29.4%), glycine (G: 44.6%), serine (S: 12.1%), tyrosine (Y: 5.2%), valine (V: 2.2%) and acidic amino acids (aspartic acid, 1.3%; glutamic acid, 10%) [5] with repetitive hydrophobic units (GAGAGX; X = S, V, or Y [6,7]). Actually, the exact fibre structure remains a source of debate because of the poor crystallinity of the materials. Less than 10 very broad peaks or rings are observed on the X-ray fibre diagram ∗ Corresponding author. Tel.: +33 1 4427 2785; fax: +33 1 4427 3021. E-mail addresses: aline.percot@upmc.fr, philippe.colomban@upmc.fr (A. Percot). 1 Present address: Faculty of Physics, Hanoi National University of Education, 136 Xuan Thuy, Cao Gay, Hanoi, Viet Nam. [8–10]. Two models are proposed: Silk I, for chemically treated silk and, Silk II in the excreted fibre [11–16]. Orthorhombic (P22 1 2 1 ) or monoclinic (P2 1 ) Silk II structure is expected to be made of beta sheets. However the Bragg peak width, i.e. the coherence length hardly reaches 2 (inter-chain distances) and 7 nm (along the chain), i.e. 2 or 10 times the unit cell size. Different structure types and associated mechanical properties have been recognised from their tensile stress/strain behaviour to depend on the fibre history (age, water content, thermal treatments, etc.) whatever the producing creature [17]. We will focus our attention on the formation of silk fibre from the biosynthesis in the posterior part of the gland up to the double-fibre bave expelled from the worm spinneret. The silkworm gland is a highly specialised secretory system divided into three main parts according to morphology, secretory products and other characteristics (Fig. 2): (i) the posterior part, where the fibroin is synthesised, secreted and temporarily stored, (ii) the middle part, S-shaped, that functions as a huge reservoir for the liquid silk in addition to the production of sericins, proteins which coat and cement the two fibroin fibres to form the bave used for the cocoon and (iii) the anterior part which consists of a con- duit from the storage reservoir to the spinneret at the mouthpart [18,19]. http://dx.doi.org/10.1016/j.vibspec.2014.05.004 0924-2031/© 2014 Published by Elsevier B.V.