Supporting information Automated Solid Phase Synthesis of Teichoic Acids Wouter F. J. Hogendorf, a Nico Meeuwenoord, a Herman S. Overkleeft, a Dmitri V. Filippov, a Diana Laverde, b Andrea Kropec, b Johannes Huebner, b Gijsbert A. Van der Marel *a and Jeroen D. C. Codée *a a Leiden Institute of Chemistry, Leiden University, P.O. Box 9502, 2300 RA Leiden, The Netherlands. Fax: +31-5274307 Tel: +31-5274280; E-mail: jcodee@chem.leidenuniv.nl; marel_g@chem.leidenuniv.nl b Division of Infectious Diseases, Department of Medicine, University Medical Center Freiburg, Hugstetter Strasse 55, 79106 Freiburg, Germany. Experimental section General All chemicals (Acros, Fluka, Merck, Schleicher & Schuell, Sigma-Aldrich, Genscript) were used as received and reactions were carried out dry, under an argon atmosphere, at ambient temperature, unless stated otherwise. Column chromatography was performed on Screening Devices silica gel 60 (0.040- 0.063 mm). TLC analysis was conducted on HPTLC aluminium sheets (Merck, silica gel 60, F245). Compounds were visualized by UV absorption (245 nm), by spraying with 20% H 2 SO 4 in ethanol or with a solution of (NH 4 ) 6 Mo 7 O 24 ·4H 2 O 25 g/l and (NH 4 ) 4 Ce(SO 4 ) 4 ·2 H 2 O 10 g/l, in 10% aqueous H 2 SO 4 followed by charring at +/- 140 o C. Some unsaturated compounds were visualized by spraying with a solution of KMnO 4 (2%) and K 2 CO 3 (1%) in water. Optical rotation measurements ([α] D 20 ) were performed on a Propol automated polarimeter (Sodium D-line, λ = 589 nm) with a concentration of 10 mg/ml (c = 1), unless stated otherwise. Infrared spectra were recorded on a Shimadzu FT-IR 8300. 31 P, 1 H, and 13 C NMR spectra were recorded with a Bruker AV 400 (161.7, 400 and 125 MHz respectively) or a Bruker DMX 600 (600 and 150 MHz respectively). NMR spectra were recorded in CDCl 3 with chemical shift (δ) relative to tetramethylsilane, unless stated otherwise. When D 2 O was used, 1 H-NMR spectra were recorded with chemical shift relative (δ) to HDO (4.755 ppm), 31 P spectra were measured with chemical shift relative to 85 % H 3 PO 4 (external standard) and 13 C-NMR spectra were recorded with chemical shift relative to TMS (external standard). High resolution mass spectra (HRMS) were recorded by direct injection (2 µl of a 2 µM solution in water/acetonitrile; 50/50; v/v and either 0.1 % formic acid or 10mM ammonium formate for the oligomers) on a mass spectrometer (Thermo Finnigan LTQ Orbitrap) equipped with an electrospray ion source in positive mode (source voltage 3.5 kV, sheath gas flow 10, capillary temperature 275 o C) with resolution R = 60000 at m/z 400 (mass range m/z = 150-2000) and dioctylphthalate (m/z = 391.28428) as a lock mass. The high resolution mass spectrometer was calibrated prior to measurements with a calibration mixture (Thermo Finnigan). LC- MS analysis was performed Procedure for automated solid-phase synthesis, purification and global deprotection of TA oligomers Aminopropyl modified controlled pore glass support (CPG, Fluka) was loaded with (glucosyl)glycerol succinates 1b or 2b and the loading was determined (loading: 100 mol/g CPG) using the method described by Pon. 1 The automated syntheses were performed on a synthesizer (ӒKTA oligopilot plus, GE Healthcare) on a scale of 100-150 mg functionalized CPG (10-15 mol glycerol derivative) and started off with acidolysis of the dimethoxytrityl ether using 3 % dichloroacetic acid in toluene (15 ml, 3 min). After flushing with acetonitrile (5ml, 1 min), the resulting alcohol was reacted with phosphoramidites 1a or 2a (0.1 M in ACN, 5 eq) and 5-benzylthiotetrazole (BTT, 0.3M in acetonitrile, 22.5 eq) for 5 min using a cycled flow. After flushing with acetonitrile (5ml, 1 min), oxidation of the intermediate phosphite was performed using I 2 (0.05 M in pyridine/H 2 O 9/1, 2 ml, 1 min). A flushing step with acetonitrile (5ml, 1 min) was followed by a capping step (1 ml of a 1/1 mixture of capping solution A (20 v/v % N-methylimidazole in acetonitrile) and capping solution B (20 v/v % Ac 2 O, 15 v/v % 2,6-lutidine in acetonitrile for 12s). After flushing with acetonitrile (5ml, 1 min), a detritylation step was performed using the before mentioned cocktail and the molecule was elongated using phosphoramidites 1a or 2a using the same set of reactions (coupling, oxidation, capping, detritylation). The average coupling efficiency was measured by quantitative UV-detection (400 nm) of the dimethoxytrityl cation during each detritylation step. When the desired length was obtained, spacer phosphoramidite 3 (0.1 M in ACN, 2 x 5 eq, 2 x 5 min) was coupled to the CPG-TA-oligomer using Electronic Supplementary Material (ESI) for Chemical Communications This journal is © The Royal Society of Chemistry 2011