Fullerene-like amorphous carbon nitride film surface properties and evaluating the initial adsorption kinetics of albumin and fibrinogen

The authors report a PTFE contact angle of 107.3 ± 2.2° from their home-built setup, and state it falls within the measured range (108.7 ± 2.5°) of a commercially available contact angle instrument — the Dropometer. This is the paper's only use of the Dropometer, and it is the reference side of the comparison.

Water contact angles reported in Table 1 are 72.0 (1.5)° (0% N₂), 58.3 (3.3)° (10% N₂), 68.0 (1.0)° (20% N₂), and 70.2 (3.4)° (30% N₂). The trend is not monotonic: wettability improves sharply at 10% N₂ and then falls back at 20% and 30%, ending close to the nitrogen-free film.

The paper's Conclusions: "Contact angle measurements showed that the incorporation of nitrogen into the FL-CNx films improved the hydrophilic surface properties, with the FL-CNx-10 film showing the best wettability."

Figure 4a compares ka kinetics for fibrinogen and human serum albumin with the film surface wettability across the %N₂ plasma discharge gas series. Figure 4b makes the same comparison against the Raman I(D)/I(G) ratio. The kinetics themselves were measured by surface plasmon resonance, not by contact angle.

Introducing higher at% N into the films (FL-CNx-10 and FL-CNx-20) reduced ka values by an order of magnitude for both HSA and fibrinogen — but the paper's next sentence reverses it: "However, this trend reverses and the ka values for HSA and Fib increase an order of magnitude for the FL-CNx-30 film." The effect is non-monotonic in nitrogen content, in the same way the contact angle is.

The paper's stated conclusion, after comparing binding kinetics against wettability, roughness, sp² clustering and atomic composition in turn: "There is no definitive property within the films that correlates with the changes in protein binding kinetics. It is most likely a combination of different surface compositions which include the amount of sp² clustering as well as the different nitrogen and oxygen functional groups at the surface." In the Results the authors write that "there is some type of correlation with the changes in the FL-CNx film properties, though it is not very obvious and clear-cut." Contact angle was one of the properties compared, as was AFM roughness, which the authors note should not play a significant role because R(RMS) values were very similar across the four films.

Association rate constants from BiacoreX SPR, fit to a 1:1 Langmuir model (Table 2): fibrinogen ka ranged from 18.8 to 203 × 10⁴ M⁻¹s⁻¹ against 2.7 to 9.2 × 10⁴ M⁻¹s⁻¹ for albumin — roughly 6- to 22-fold faster depending on film. Dissociation rates were comparable for both proteins across all films, so binding affinity was governed by association: "the ka plays a larger role in the variation of the protein's binding affinity to the surface." The authors caution that the 1:1 model "is quite simple, and it assumes that there is one independent site located on the surface", and that with multiple binding points per protein "the measured kinetics will be a result of the average of these different binding orientations."

On the nitrogen-free FL-CNx-00 film, 500 nmol/L fibrinogen left approximately 680 RU (680 pg mm⁻²) on the bare surface. Over a surface pre-saturated with albumin (1200 RU immobilised), the same injection produced a baseline shift of only 20 RU — a 34-fold reduction in fibrinogen surface capacity. Confocal imaging of an albumin:fibrinogen mixture at the serum-like weight ratio showed albumin fluorescence essentially constant across all four films. The authors state the limit of the experiment explicitly: "it is unclear at this time from the measurement on whether the Fib actually bound to the FL-CNx surface (pinholes in the HSA film), or whether the Fib actually bound to the surface or to the HSA layer."

The paper credits “Dropometer, Droplet Lab, Toronto, ON” as the commercially available contact angle instrument used as a reference range for verifying PTFE contact-angle measurements from the authors’ setup.

Water contact angles are reported for films prepared with 0%, 10%, 20%, and 30% N₂ plasma discharge gas, enabling a direct wettability comparison across the deposition series (Table 1).

Table 1 indicates the contact angle minimum occurred at 10% N₂ (58.3 (3.3)°), which the Conclusions describe as "the best wettability". The effect does not scale with nitrogen: 20% and 30% N₂ returned 68.0 (1.0)° and 70.2 (3.4)°, close to the nitrogen-free film's 72.0 (1.5)°. Beyond 10% N₂, more nitrogen in the plasma discharge did not give a more wettable film.

The research presents contact angle (wettability) together with SPR association kinetics (ka) for fibrinogen and HSA across the %N₂ series (Fig. 4a), discussing substantial changes in ka values with nitrogen incorporation. The paper does not claim wettability explains those changes. It reports that no single film property definitively correlated with the binding kinetics, and suggests the Fib kinetics are "possibly due to different distributions of the different nitrogen/oxygen functional groups located within the FL-CNx film."

The water contact angles identify the 10% N₂ film as the most wettable in the series. The authors note that surface roughness should not play a significant role because R(RMS) values were very similar across films (1.2–2.1 nm), and that the overall N/C and O/C atomic ratios do not necessarily play a significant role because FL-CNx-10 and FL-CNx-30 had very similar ratios. They suggest the more hydrophilic nature may reflect additional surface functional groups near the surface that may not be detected through the NEXAFS measurements, consistent with earlier EELS work on films from the same deposition apparatus that found greater oxygen near the surface than in the bulk.