The impact of zwitterionic surfactants on optode-based nanosensors via different fabrication approaches and sensing mechanisms

In this work, we explored the impact of zwitterionic surfactants, sulfobetaine 16 (SB-16) and a PEG-phospholipid conjugate (DSPE-PEG), on nanosensor performance. We fabricated four sensors (for Na+, K+, Al3+, and O2) and examined how these surfactants influenced various aspects, including fabrication methods, sensing mechanisms, and the incorporation of nanomaterials. Our results highlighted SB-16's role in enhancing selectivity in ion-exchange sensors (Na+ and K+) while maintaining sensitivity akin to its PEG counterpart. The liquid–liquid extraction based sensors (Al3+) were unaffected by surfactant choice, while the O2 sensors that operate via collisional quenching exhibited reduced sensitivity with SB-16 when compared to its PEG-based counterpart. Additionally, the SB-16 sensors proved adaptable to different fabrication approaches (SESE – single emulsion solvent evaporation and FNP – flash nanoprecipitation), showcased good cell viability and maintained a functional lifetime of at least five days. Furthermore, via the use of quantum dots, we showed that it is possible to incorporate other nanomaterials into the sensing phase of SB-16 sensors. Future investigations could target enhancing the pH stability of zwitterionic surfactants to further advance their applicability in sensor technologies.


Table S3
Sodium and potassium selectivity coefficients

Table S5
Temporal study on SB-16 potassium sensors and potassium analyte  * The QD 520 was incorporated into the optode as per the procedure by Ruckh et.al 1 .In short, in a 1.5 mL Eppendorf tube, 120 µL ethanol was added to 0.2 mg QD (80 µL of 2.5 mg/mL in THF) and was vortexed for 1 min.After the initial mix, the tube was loaded onto a centrifuge and spun for 5 mins at 8000 RCF.The supernatant was extracted and discard.80 µL of 3.3 mM dodecanthiol in THF was added to the pellet and then vortexed for 1 min to make a stock of 2.5 mg/mL QD 520 in "thiol-THF".Two of these stocks were used in one potassium optode.To evaluate if the sulphur group on the SB-16 precipitates in the presence of potassium, we studied the colloidal stability of the nanosensor under high (10 mM K + ) and low (HEPES/Tris) potassium concentrations.Post-fabrication, the nanosensors were mixed in equal parts with the analyte solutions (1.2 mL sensors: 1.2 mL analyte) and were sampled over 2 days.At the point of evaluation, the samples were extracted and prepared in triplicate for DLS as described in the methods section.As seen by the data, there was a small decrease in the particle size of the nanosensor left in HEPES/Tris (statistically significant difference, two-tailed t-test, p = .008< 0.05) but this could be attributed to the limitations of the instrument.However, the sensors in the 10 mM KCl solution showed no signs of precipitation or changes in particle size over two days indicating good colloidal stability which is in alignment with our findings in Fig. S2 (no statistically significant difference, two-tailed t-test, p = .88> 0.05).Where not visible, the error bars are smaller than that of the data points.

Fig. S5: CCK-8 Assay monitoring the viability of yeast cells under different conditions, with the absorbance monitored at 460 nm (n =3
).The assay was set-up as described by Saccomano et al 2 .As seen in graph above, the cells with SB-16 sodium sensors performed comparable to its PEG and PBS counterparts.If the SB-16 sensors were deleterious to the cells, the lowered metabolic activity would result in the decrease in the production of the orange formazan dye, resulting a lower absorbance at 460 nm.

Fig. S6: Absorbance and emission spectra of the SB-16 potassium sensors, with a quantum dot reference (n=3). A. Absorbance spectra of the potassium sensor. As analyte concentration changes, the absorbance of the CH V in the sensor also changes thus gating the emission of the quantum dot at 510 nm. B.
As the concentration of the analyte increases, the absorbance at 510 nm increases, which results in a decrease in the QD emission at 510 nm.C. The CH V emission at 710 nm is opposite to that of the QD 520 response and is linked to the absorption at 550 nm.As result, when the analyte concentration increases, the emission at 710 nm to also increase.

Fig. S7: TEM images of quantum dots (dark, opaque dots) within potassium nanosensors (larger, translucent grey spheroids).
The TEM images show that most nanoparticles are around 150 nm, which is around the ~180 nm reported by the DLS results.Some of the shapes are rather irregular/non-spherical, thus could be due to some of the nanosensors falling apart when loaded onto the TEM lacey carbon grid.

Fig. S8:
Retentability of the QD 520 in the SB16 potassium nanosensors (n=3).To do this, the sensors were concentrated via amicon filters (30 kDa) before the retentate was raised up to the original concentration (equal to that of the stock).The retentate was measured against the flow-through, stock and a HEPES/Tris control.The QD 520 channel retentate showed a 73 % signal retention, while the CH V channel showed a 93% signal retention.The lowered QD 520 signal could be attributed to some of the nanoparticles being caught in the filter.
Fig. S1Surfactant chemical structuresFig.S2Potassium nanosensor response kinetics over 18 hoursFig.S3Ionophore-free nanosensor responseFig.S4Functional lifetime of SB-16 sensors Fig. S5 Measuring cell viability response of DSPE-PEG & SB-16 sensors Fig. S6 Absorption spectra of sodium sensors (PEG and SB-16) under different analyte conditions Fig. S7 TEM images of quantum dots in SESE SB-16 sensors Fig. S8 Rentability of the QD 520 in the SB16 potassium Fig. S9 Response of the aluminum sensors to aluminum analyte Fig. S10 Effect of increasing NaBARF on the response of the aluminum nanosensors Fig. S11 Emission spectra of the (DSPE-PEG and SB-16) SESE oxygen sensors Fig. S12 Calibration curve and emission spectra for oxygen sensors fabricated with FNP Fig. S13 Stern-Volmer response of oxygen sensors fabricated via SESE Fig. S14 Absorption spectra of sodium sensors (PEG and SB-16) under different analyte conditions

Fig. S9 :
Fig. S9:Response of the Aluminum sensors to analyte ranging from 1 µM to 0.1 mM (n =3).From this calibration curve, the linear response was generated by evaluating the sensor luminescence for analyte ranging from 1 µM to 50 µM.Where not visible, the error bars are smaller than that of the data points.

Fig. S10 :
Fig. S10: Effect of increasing NaBARF on the response of the aluminum nanosensors (n=3).A. shows the luminescence response of the DSPE-PEG and SB-16 sensors with 2 mg of NaBARF in the optode.B. and C.shows the response for 3 mg and 4 mg of NaBARF in the optode.

Fig. S11 :Fig. S12 :
Fig. S11: Emission spectra of the SESE oxygen sensors in response to different dissolved oxygen concentrations (n=3).A. Emission spectra of the DSPE-PEG based oxygen sensors and B. SB-16 based oxygen sensors.As shown in Figure3but the pseudo-Stern-Volmers, the DSPE-PEG sensors have a higher sensitivity than its' SB-16 counterpart.Where not visible, error bars are smaller than that of the data points.

Fig. S14 :
Fig. S14: Absorption spectra of the sodium sensors under different analyte conditions (n=3). A. Absorption spectra of the DSPE-PEG Na sensors under various sodium dilutions (buffered in HEPES/Tris at pH 7.4) and acid and based points.As expected, the chromoionophore is fully protonated at 650 nm under acid conditions (red line) and deprotonated at 650 nm under basic conditions.B. Absorption spectra of the SB-16 Na sensors under the aforementioned conditions.However, at 650 nm in an acidic environment, the SB-16 sensors seem to have nearly twice the absorbance of its PEG counterpart, thus indicating the sensor might not be completely stable under extreme pH variations.