Dd2 600 mhz nmr spectrometer

All chemicals were purchased from commercial suppliers and used without further purification. NMR spectra of all synthesized compounds were carried out on an Agilent DD2-400 MHz NMR or Agilent DD2-600 MHz NMR spectrometer with ProbeOne. All chemical shifts of ¹H and ¹³C signals were reported in parts per million using TMS as internal standard at 25 °C and all spectra were calibrated using the respective solvent signal. Mass spectra were recorded on an Advion ExpressIon CMS or Xevo T-QS (Waters) using electrospray ionization. TLC analyses for reaction control were performed on Merck Silica Gel 60 F₂₅₄ TLC plates and visualized using UV light. Analytical HPLC was performed on VWR Hitachi with an Agilent C18 column (Agilent Zorbax 300SB-C18, 100 mm × 4.6 mm) and acetonitrile/water (0.1% TFA each) as mobile phase. Chromatographic separations were performed using automated flash column chromatography on Isolera Four (Biotage) using silica gel cartridges (SNAP KP-Sil; 10 g or 25 g) and reversed phase HPLC system Knauer Azura with Zorbax 300SB-C18 semi-preparative column and acetonitrile/water (0.1% TFA each) as mobile phase. Starting materials such as anhydrous solvents, dimethyl 4-hydroxypyridine-2,6-dicarboxylate (TCI Europe) and 4,13-diaza-18-crown-6 (Merck) were purchased and used as received. Methyl 6-(chloromethyl)picolinate was prepared according to the literature [44 (link)].

³¹P-NMR measurements were performed as described earlier^{29 (link)}. In detail, spectra were recorded on DD2 600 MHz NMR spectrometer (Agilent) using a OneNMR probe and Avance Neo 600 MHz NMR spectrometer (Bruker) using a BBFO probe, both tuned at the resonance frequency of the ³¹P nucleus, with 5 mm outer diameter tubes containing about 1.2 ml thylakoid suspension at a Chl content of about 8–10 mg/ml. As tested earlier, at this concentration, no magnetic orientation of the membranes occurs^{27 (link)}. The temperature was controlled within 0.1 °C; spectra were recorded using a 40°rf pulse, an interpulse time of 0.5 s and no ¹H-decoupling. ³¹P chemical shifts are reported relative to 85% H₃PO₄ in water (δP = 0 ppm) used as an external reference. Spectral deconvolutions were performed by using the DMfit software^{70 (link)}.

HPLC chromatograms were acquired using the Waters 2695 Alliance liquid chromatographic system (Milford, MA, USA), equipped with the UV Waters 2996 photodiode array detector (Milford, MA, USA). Empower3 software (Waters, Milford, MA, USA) was used to record and process the signals.
1D ¹H and ¹H-DOSY NMR spectra were collected on an Agilent Technologies DD2 600 MHz NMR spectrometer (Santa Clara, CA, USA), equipped with the HCN Cold probe.

All NMR spectra were obtained with Agilent Technologies DD2 600 MHz NMR spectrometer at 5, 25, and 37 °C using a triple resonance cold probe. Standard 1D ¹H spectra were acquired with the use of DPFGSE, watergate 3919 or PRESAT solvent suppression. Diffusion coefficient measurements were performed by a spin-echo pulse sequence with PFG gradient strengths between 0.49 and 29.06 G cm⁻¹. NOESY spectra were acquired with mixing times of 80 and 150 ms. Assignment of imino protons of guanine residues was done by 1D ¹⁵N-edited HSQC experiments performed on residue specific 10% ¹⁵N, ¹³C-isotopically labeled oligonucleotides. Measurements at different pH were performed at pH 4.7, 6.0 and 7.2. Measurements under molecular crowding conditions were performed using 40% w/v dPEG (8000 MW) (Polymer Source Inc., Dorval, Canada) in water 7 days after annealing. KH₂PO₄ was used in experiments with 100 mM K⁺ ions. NMR spectra were processed and analyzed using VNMRJ (Varian Inc.) and Sparky (UCSF) software.

Deuterium NMR studies were conducted on an Agilent DD2 600 MHz NMR spectrometer equipped with 3.2 mm MAS triple resonance magic-angle spinning probe, without spinning. Static deuterium NMR spectra were recorded at 92.17 MHz using a standard quadrupolar echo sequence with π/2 pulses of 4 μs and an inter-pulse delay of 50 μs. Temperature was regulated to 30 °C and 128,000 acquisitions were typically acquired. Deuterium spectra were referenced externally to D₂O. Prior to Fourier transformation the deuterium free induction decay was left-shifted to the top of the echo and 500 Hz line-broadening applied. All NMR data were processed in matNMR53 .