Scientific Protocol for Preparation and Characterization of Single-Walled Carbon Nanotubes

Key Insight: Chirality-pure, colloidally stable SWCNTs in various surfactant environments provide a robust platform for investigating nanoscale opto-thermal effects.

Article Title: Advanced Preparation and Multimodal Spectroscopy of Single-Walled Carbon Nanotubes (SWCNTs)

Preparation of SWCNTs

All chemicals were purchased from Sigma Aldrich (Germany) unless stated otherwise. Experiments were performed with (6,5)-enriched SWCNTs (Sigma Aldrich, Signis SG65i, CoMoCAT synthesis technology).

• A mixture was prepared containing 150 µl of 2 mg ml⁻¹ single-stranded DNA (e.g., (GT)₁₀) in 1× PBS buffer (pH 7.4), 75 µl of 2 mg ml⁻¹ SWCNT in PBS, and 75 µl PBS.
• This was followed by tip sonication (Fisher Scientific, FB120, 120 W, amplitude 35%, 9 s pulse on, 1 s off, 15 min).
• Centrifugation at 21,000g for 30 min was performed; the supernatant was collected. This procedure was repeated two more times.
• The final supernatant was stored at 4 °C.
• For (GT)₁₀-SWCNT experiments in D₂O, PBS buffer was prepared with D₂O.

• (DOC)-SWCNTs were mixed with PEG (6 kDa, 8% w/v), dextran (70 kDa, 4% w/v), DOC (0.025% w/v), SDS (0.5% w/w), and SC (0.5%-0.9% w/w in 0.1% increments).
• Chiralities were adjusted by adding HCl. A one-step approach used specific volumes of HCl and NaClO (10-15% available chlorine) for pH-driven and electronic separation, yielding monochiral (6,4)-SWCNTs in the bottom phase (B3).
• The solution was dialysed (300 kDa bag) against 1% DOC solution to remove dextran, resulting in a stable 1% DOC-(6,4)-SWCNT solution.

• A standard mixing protocol was used: 150 µl of 2% (m/v) DOC in H₂O was mixed with 150 µl of 2 mg ml⁻¹ SWCNTs in H₂O.
• Tip sonication and centrifugation were performed as for (GT)₁₀-SWCNTs; the supernatant was stored at 4 °C.
• SDBS- and SC-functionalized SWCNTs were prepared identically.

• A precise chemical reaction was induced: 20 µl of 4 mM 4-nitrobenzol diazonium tetrafluoroborate was added to 20 ml of 10 nM SDBS-SWCNTs.
• The mixture was irradiated with green light (550 nm) while stirring for 15 min.
• The solution was then mixed with an equal volume of acetonitrile (ACN); SWCNTs precipitated and the pellet was washed with H₂O two to three times to remove residual SDBS and ACN.
• The precipitate was redispersed in 1% DOC by tip sonication (15 min) and centrifugation (30 min at 21,000g); the supernatant was collected. The resulting SWCNT length was about 600 nm.

• Confirmation of stability was achieved using absorbance, 1D and 2D fluorescence spectroscopy, and atomic force microscopy (average length ~600 nm).
• Chirality-pure (6,5)- and (6,4)-SWCNTs were prepared to exclude impurity effects.

NIR Spectroscopy

• Measurements were conducted on 0.5 nM (GT)₁₀-SWCNTs, with or without analytes (2 µM riboflavin, 100 µM ascorbic acid), or 0.5 nM DOC-, SC-, SDBS-SWCNTs, using an Olympus IX73 microscope and a 561 nm solid-state laser.
• Emission spectra were captured with an Andor iDus InGaAs 491 array NIR detector and a Shamrock 193i spectrometer.

• The same setup was used as for 1D, with tunable excitation via a monochromator (MSH150) and an LSE341 light source.
• Samples included 2 nM SWCNTs in various surfactants and (GT)₁₀-SWCNTs in PBS (D₂O) at pH 7.4.

FCS Measurements

• A MicroTime 200 system (PicoQuant) was employed, featuring pulsed lasers at 485 nm and 530 nm, an Olympus IX73 confocal microscope (60× water objective, NA 1.2), and SPAD detectors.
• The focus was on (6,5)-SWCNTs due to detector sensitivity. 1 nM samples were excited at 485 nm (40 MHz) or 532 nm (CW for DOC-(6,4)-SWCNTs).
• Light passed through a dichroic mirror, a 900-nm long-pass filter (800 nm for DOC-(6,4)-SWCNTs), a 50-µm pinhole, and was directed to detectors.
• Refractive index and viscosity corrections were applied via collar settings.

• The standard correlation function was defined as G(τ) = ⟨I(t)I(t+τ)⟩ / ⟨I(t)²⟩.
• Diffusion correlation was described by G_D(τ) = (1/N)[1+τ/τ_D]⁻¹[1+τ/(w²τ_D)]⁻¹/².
• The diffusion constant D = w_xy²/(4τ_D) and structural parameter w = w_z/w_xy were calibrated using Atto 488 dye (1 nM, D_t = 400 μm² s⁻¹); the excitation volume was 1.5 fl.
• The fitting equation in Igor Pro 6.34A included a dark state term.

• Controls ruled out artifacts: Atto 488 control showed a slight decrease in G(0) but no change in normalized autocorrelation or diffusion time, ruling out sample heating.
• Temperature was constant at 10 μW and 90 μW.
• Brightness of (GT)₁₀-SWCNTs increased linearly with laser power, with no non-linear effects.
• The apparent particle number increased with power (3 to 9.3) due to low quantum yield.
• Diffusion behavior was reversibly switched by changing excitation power.
• Chirality-purified (GT)₁₀-(6,5)-SWCNTs showed similar diffusion; sample purity was high.
• CW excitation gave the same results as pulsed; diffusion on the ms timescale was less affected by excitation timing.
• Absorbance spectra were unchanged with analytes, indicating no aggregation or dissociation.
• In 80% glycerol/water, almost no diffusion changes were observed; in 20% glycerol/water, power-dependent changes were similar to water.
• Simulations (random walk) qualitatively confirmed that excitation-induced diffusion changes lead to power-dependent autocorrelation functions.

THz Measurements

• A two-colour air plasma filament generated broadband THz probe pulses from 50 fs, 800 nm laser pulses.
• A 400 nm optical pump (frequency-doubled) was used. Changes in THz absorption were measured as a function of pump–probe delay (0.25 ps to 300 ps).
• A windowless free-flowing jet (20 μm thickness) was used; 80 ml of ~100 nM SWCNT solution was circulated for 96 h; a defoaming agent was added.
• The THz field was detected via electro-optic sampling with a 100 µm GaP crystal.
• The difference in THz transmission was expressed as ΔmOD; positive values indicate decreased transmission upon optical excitation.
• Fluence was varied: 50, 120, 200 mJ cm⁻². Data is shown for 200 mJ cm⁻² unless stated.

Wide-field Tracking of SWCNTs

• A thin film spacer (2.3 µm Mylar) was placed between glass slides; 50 µl of 0.1 nM purified DOC-SWCNT was added.
• A 561 nm laser (200 mW, 100 W cm⁻²) was coupled to an Olympus IX73 microscope with a 100× oil-immersion objective (NA 1.35).
• NIR imaging was performed with an InGaAs camera (Cheetah, Xenics 640, 640×512 pixels, thermoelectrically cooled).
• A dichroic mirror and 900 nm long-pass filter were used; images were captured at 7 fps, 140 ms exposure.
• Data analysis was performed with Python 3.10.5; trackpy identified bright spots; traces >100 frames were analyzed.
• Ensemble time-averaged mean-squared displacement (MSD) was computed.

Computation of Friction and Diffusion in Water

• LAMMPS was used to model a graphene slab (2.5×2.6 nm², 1,600 water molecules) and a (6,5)-SWCNT (3×3×4.1 nm³, 1,100 water molecules).
• Both non-polarizable and polarizable systems were simulated with CVFF and IFF-CVFF parameters.
• Non-polarizable: neutral C with LJ interactions. Polarizable: each C with two flexible negatively charged dummy atoms (π-orbitals) via harmonic bonds and angles.
• The polarizable model includes Coulombic interaction with water.

• The friction coefficient was calculated as λ_GK = (1/(A n k_B T)) ∫⟨FL(t)FL(0)⟩ dt.
• For non-polarizable graphene: ~2×10⁴ N s m⁻³; polarizable: ~6.5×10⁴ N s m⁻³ (agreement with ab initio estimates).
• For the SWCNT external surface: non-polarizable 6.5×10⁴ N s m⁻³, polarizable 15×10⁴ N s m⁻³.
• The IFF-CVFF polarizable model was validated against bulk and interfacial properties.

• A (6,5)-SWCNT (length 4.1 nm) was placed in a 3D periodic box of 140,000 water molecules (flexible SPC).
• After NPT equilibration, a 20 ns trajectory was run (0.5 fs timestep, snapshot every 1 ps).
• The SWCNT was end-capped with hydrogen and allowed to diffuse unconstrained (NPT, 298 K, 1 atm).
• The MSD of the center-of-mass was computed; D = slope/6.
• The excited state was modeled by adding an axial dipole: charges of 44 virtual atoms were modified by ±0.005 e (total 0.22 e) in the polarizable case; or, for non-polarizable, two rings of 22 C atoms with ±0.01 e each.
• A dipole-free excitation was also investigated (outer rings −0.005 e, central ring +0.01 e; negligible dipole moment).