1030 Research Article Received: 12 February 2008 Revised: 20 June 2008 Accepted: 25 July 2008 Published online in Wiley Interscience: 10 September 2008 (www.interscience.com) DOI 10.1002/mrc.2311 Spectral assignments and anisotropy data of cellulose I α : 13 C-NMR chemical shift data of cellulose I α determined by INADEQUATE and RAI techniques applied to uniformly 13 C-labeled bacterial celluloses of different Gluconacetobacter xylinus strains Stephanie Hesse-Ertelt, a* Raiker Witter, b Anne S. Ulrich, b,c Tetsuo Kondo d and Thomas Heinze a* Solid-state 13 C-NMR spectroscopy was used to characterize native cellulose pellicles from two strains of Gluconacetobacter xylinus (ATCC 53582, ATCC 23769), which had been statically cultivated in Hestrin–Schramm (HS) medium containing fully 13 C-labeled β -D-glucose-U- 13 C 6 as the sole source of carbon. For both samples, the 13 C-NMR chemical shifts were completely assigned for each 13 C-labeled site of cellulose I α with the aid of 2D refocused INADEQUATE NMR. To determine the principal chemical shift tensor components, a pulse sequence based on the recoupling of anisotropy information (RAI) was applied at 10 kHz MAS. The detailed 13 C tensors of cellulose I α from different bacterial celluloses are thus available now for the first time, and these results have been compared with previously published data of nonenriched material and with theoretical predictions. Copyright c  2008 John Wiley & Sons, Ltd. Keywords: NMR; 13 C-NMR; bacterial cellulose; Gluconacetobacter xylinus; Acetobacter xylinum; recoupling of anisotropy information; RAI; INADEQUATE Introduction Solid-state NMR spectroscopy has been widely used to study the structure of cellulose from different sources such as algae, plants, and bacteria, and of its derivatives. Cellulose consists of 1–4 linked β -D-glucopyranose repeating units which form fibrous structures with regions of high crystallinity as well as amorphous parts. Elucidation of the crystal structure of native cellulose still remains one of the most challenging tasks in the field of cellulose research. Using cross-polarized/magic-angle spinning (CP/MAS) 13 C{ 1 H}-NMR on highly crystalline native cellulose I, VanderHart and Atalla [1] demonstrated the presence of two allomorphs, I α and I β . It was shown that the I α /I β ratio in native cellulose differs greatly from species to species. Cellulose I α is the dominant form in algae and bacterial celluloses, while I β is dominant in higher plants and animal celluloses. [2–5] The first 13 C-NMR resonance assignment was published for carbons C1, C4, and C6, and the cluster of signals between 70 and 80 ppm was attributed to C2, C3, and C5. [1] These conclusions were subsequently confirmed by solid- state INADEQUATE NMR. [6] Further details based on correlation spectroscopy were presented by Sakellariou et al. [7] and Cadars et al., [8] and the three-dimensional cellulose structure and NMR investigations thereof were discussed by Sternberg et al. [9] and Witter et al. [10] The complete assignment of all carbon sites in the crystalline structure was finally possible using specifically 13 C- labeled D-glucose and D-glycerol for biosynthesis, [11,12] allowing to resolve the individual chemical shifts of C2, C3, and C5. Next, the structural assignment of all 13 C-NMR signals in the two different anhydroglucose rings were succeeded by solid-state INADEQUATE NMR of specifically prepared allomorphs I α (purified Cladophora) and I β (purified tunicate cellulose). [13] It was clearly demonstrated that both I α and I β phases contain two magnetically different D-glucose residues in the unit cells. Slightly different isotropic chemical shift values were found by Jaeger et al., [14] who assigned all carbon sites in unpurified, uniformly 13 C-enriched bacterial cellulose (BC) of the strain Gluconacetobacter xylinus (previously ∗ Correspondence to: Stephanie Hesse-Ertelt and Thomas Heinze, Centre of Excellence for Polysaccharide Research, Friedrich Schiller University of Jena, Humboldtstrasse 10, 07743 Jena, Germany. E-mail: stephanie.hesse@uni- jena.de; thomas.heinze@uni-jena.de a Centre of Excellence for Polysaccharide Research, Friedrich Schiller University of Jena, Humboldtstrasse 10, D-07743 Jena, Germany b KIT, Institute of Biological Interfaces, Forschungszentrum Karlsruhe, 76021 Karlsruhe, Germany c KIT, Institute of Organic Chemistry, University of Karlsruhe, Fritz-Haber-Weg 6, 76131 Karlsruhe, Germany d Bio-architecture Center (KBAC) and Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan Magn. Reson. Chem. 2008, 46, 1030–1036 Copyright c  2008 John Wiley & Sons, Ltd.